Detection, quantification, and significance of basic fibroblast growth factor in the aqueous humor of man, cat, dog and pig. By using a highly specific and sensitive heterologous radioimmunoassay, we determined the content of basic fibroblast growth factor (bFGF) in fresh samples of aqueous humor obtained from human (n = 18), feline (n = 8), canine (n = 12), and porcine (n = 10) eyes by paracentesis. The content of bFGF in human aqueous humor ranged from 0.480 to 1.44 ng ml-1 (mean +/- S.D. = 1.074 +/- 0.297 ng ml-1); in feline samples, from 0.672 to 1.152 ng ml-1 (1.016 +/- 0.158 ng ml-1); in canine samples, from 0.640 to 1.232 ng ml-1 (1.026 +/- 0.171 ng ml-1); and in porcine samples, from 0.627 to 0.957 ng ml-1 (0.728 +/- 0.115 ng ml-1). These values were confirmed by means of a dot/slot-blot technique. For all species, the aqueous samples had normal protein levels that ranged from 5 to 19 mg dl-1. There was no correlation of the content of bFGF with the level of protein or with age of the human subjects. The similarity in the concentrations of bFGF in the aqueous humor as well as the stability of the blood-aqueous barriers of all four species indicate that cats, dogs, and pigs can serve as suitable animal models for the study of the role of bFGF in health and disease. We suggest the possible involvement of bFGF in the pathogenesis of anterior-segment disorders, such as neovascular glaucoma, and in the wound-healing response of limbal tissues after glaucoma filtration surgery.


Immunohistochemical localization of hyaluronan synthase in cornea and conjunctive of cynomolgus monkey. Distribution of hyaluronan synthase was investigated in cornea and conjunctiva of Cynomolgus monkeys (Macaca fascicularis) using polyclonal antibodies against the streptococcal enzyme. Strong immunoreaction was found in the cell membranes of the corneal endothelium, corneal epithelium, and most of the conjunctival epithelium. In the corneal epithelium all cells except the basal ones stained. In the conjunctiva all cylindrical cells stained, whereas among the goblet cells one type showed intense membrane staining, the other remained unstained. In the limbal portion of the conjunctival epithelium, which in many other respects differs morphologically and functionally from the remaining conjunctiva, all membranes of the different layers of the stratified epithelium except the most superficial ones, appeared unstained. Staining was also seen in all stromal fibroblasts and capillary endothelial cells.


Description of an acylpeptide hydrolase from lens. Acylpeptide hydrolase, which catalyses the hydrolysis of blocked N-terminal amino acids from peptide substrates, has been identified in the extracts from beef, human, rabbit and rat lens. In bovine lens sections, lower activity was observed in nuclear and inner cortical regions compared to the outer cortical region. The enzyme from bovine lens showed a high molecular weight nature, eluting between alpha and beta crystallins during Sephadex G-200 chromatography. The activity has a pH optimum around 7.8 when assayed with N-acetyl-Ala-p-NA as substrate. The enzyme was capable of hydrolyzing a variety of blocked peptides including N-acetyl-(Ala)2, Me-O-Suc-Ala-Ala-Pro-Val-p-NA, N-Acetyl-Met-Leu-Phe, Acetyl-Ser-Gln-Asn-Tyr and N-formyl-Met-p-NA. In each case the enzyme released an N-blocked amino acid and exposed a free amino group as judged by thin layer chromatography. Neither Ala-p-NA nor N-acetyl-Ala were hydrolysed by the same enzyme preparation. The enzyme activity from human and bovine lens was completely inhibited by DFP, and partially inhibited by PMSF, penicillin-G and ampicillin. These preliminary results show that lens tissue has an active acylpeptide hydrolase, however, a partially purified enzyme preparations was not able to cleave the acetyl-Met- from native alpha A-crystallin in vitro suggesting that the N-terminus of native crystallins is not accessible to the enzymes.


Proenkephalin A derivatives in lacrimal gland: occurrence and regulation of lacrimal function. The derivatives of proenkephalin A were measured in acid extracts of rat lacrimal glands by specific radioimmunoassay. Glands from adult male rats contained all four derivatives of the opiate precursor. The content in the gland of proenkephalin A-derived peptides Met5-enkephalin, Leu5-enkephalin, Met5-enkephalin Arg6-Phe7 and Met5-enkephalin Arg6-Gly7-Leu8 indicates tissue specific processing with an enhancement of the heptapeptide. The effect of enkephalins on the activity of adenylate cyclase in lacrimal membranes was measured and compared with the effect of the synthetic enkephalin analogue D-ala2-methionine enkephalinamide (DALA) that inhibits both lacrimal protein secretion and lacrimal adenylate cyclase. In the presence of the peptidase inhibitors thiorphan and bestatin, the inhibition of forskolin-stimulated adenylate cyclase activity by Met5-enk, Leu5-enk, Met5-enk Arg6-Phe7 and DALA were identical. Maximum inhibition was approximately 35% at a dose of 50 microM enkephalin. Addition of the octapeptide, Met5-enk Arg6-Gly7-Leu8 resulted in decreased adenylate cyclase activity; however, the effect was not statistically significant. Activation of delta opioid receptors by the endogenous enkephalins is indicated by the reversal of adenylate cyclase inhibition in the presence of the delta-receptor antagonist ICI 174864. The data support the physiological significance of in vitro inhibition of lacrimal secretion by DALA and indicate a possible role for endogenous enkephalins in lacrimal function.


The phagocytosis of ROS by RPE cells is inhibited by an antiserum to rat RPE cell plasma membranes. A polyclonal antiserum to a rat retinal pigment epithelium (RPE) plasma membrane-enriched fraction has been utilized to identify candidate receptor proteins which may be involved in the phagocytosis of rod outer segments (ROS) by the RPE. Immunoblots of RPE cell extracts show that the the antiserum recognizes a number of glycoproteins, including two with M(r)s of 174 and 75 kDa. The antiserum also recognizes their non-glycosylated counterparts, with M(r)s of 169 and 65 kDa, respectively, which are synthesized after treatment of the cells with tunicamycin B2. Immuno-precipitation of [35S]-methionine-labeled RPE cell extracts also demonstrates the presence of antibodies to these same glycoproteins as well as to other proteins. The antiserum inhibits the binding of ROS to the RPE, which subsequently results in a decrease in the ingestion of ROS. ROS phagocytosis by the RPE is inhibited by 97% in the presence of a 1:10 dilution of the IgG fraction of the antiserum. Phagocytosis recovers to normal levels after 4-6 hr of chase in the absence of antibodies. After sequential adsorption of the IgG fraction to monolayers of fixed RPE cells, which removes RPE surface-specific IgGs, the extent of inhibition of ROS phagocytosis produced by the IgG fraction is reduced. Using immunoblotting we have identified a number of surface-specific immunoreactive bands which are adsorbed out of the antiserum, including the 174 and 75 kDa bands. These data give further support to the hypothesis that ROS phagocytosis is a receptor-mediated process, which occurs via specific cell surface glycoprotein receptors.


Immunocytochemical study of extracellular matrix components during lens and neural retina regeneration in the adult newt. We have conducted an immunocytochemical study of fibronectin, laminin, heparan sulfate proteoglycans and nidogen-entactin during lens and neural retina regeneration in the adult newt from 0 to 60 days. In the normal eye, fibronectin was detected in the corneal stroma and Descemet's membrane, in dorsal and ventral irises and lens capsule but not in Bowman's membrane of the cornea. In normal neural retina, fibronectin was found in Bruch's and inner limiting membranes. Heparan sulfate proteoglycans gave a slight signal in both irises and the lens capsule. Nidogen-entactin distribution in the cornea was similar to that of fibronectin; it was absent from the stroma of both irises, and the signal was weak in the pigmented iris epithelium. Nidogen-entactin was not detected in the lens capsule and inner limiting membrane of the neural retina but was present in Bruch's membrane. During the first 15 days of lens regeneration, fibronectin and nidogen-entactin decreased but did not disappear from the pupillary margin of both irises, and no signal was obtained for laminin and heparan sulfate proteoglycans. From day 15 to day 60 fibronectin and nidogen-entactin increased in both irises and lens capsule. The signal for laminin was restricted to the lens capsule. Heparan sulfate proteoglycans gave a slight signal in both irises and in the lens capsule. During the first 25 days of neural retina regeneration, fibronectin was the first to appear in Bruch's membrane and the cell border of the new neuroepithelium and remained during the entire process. Laminin appeared after 41 days in the inner limiting and Bruch's membranes, but by day 50 it appeared as a weak signal only in the inner limiting membrane. Heparan sulfate proteoglycans were not detected at any of the regeneration stages studied. Nidogen-entactin was only detected in Bruch's membrane and around the cells and blood vessels of the new neural retina. Later it was detected in the inner limiting membrane but not in Bruch's membrane. Thus, the results obtained showed that extracellular matrix components do change during both lens and neural retina regeneration. These changes may play an important role during both regenerating processes.


Pressures in the juxtacanalicular tissue and Schlemm's canal in monkeys. A micropuncture technique involving the use of microcannulas with tip diameters less than 5 microns was used to measure the pressure in Schlemm's canal and in the meshwork at distances approximately 7 and 14 microns from the inner wall of Schlemm's canal. In one set of experiments where the spontaneous intraocular pressure (IOP) was 12.2 +/- 0.5 cmH2O and the Schlemm's canal pressure (PSc) was 7.6 +/- 0.7 cmH2O, the pressure at 7 microns from the inner wall of Schlemm's canal was found to be 8.9 +/- 0.7 cmH2O and at a distance of 14 microns, 11.0 +/- 0.5 cmH2O--that is, 1.3 +/- 0.2 and 3.4 +/- 0.3 cmH2O respectively, higher than the PSc. In another set of experiments, the spontaneous IOP and PSc were also measured and then the IOP was increased by means of an external reservoir and measured once again. Spontaneous IOP was 16.0 +/- 1.3 cmH2O and the PSc was 11.5 +/- 1.4 cmH2O before the IOP was increased. After the IOP was increased to 20.2 +/- 1.2 cmH2O, the PSc was 11.7 +/- 1.6 cmH2O. When the microcannula was introduced into the juxtacanalicular tissue to locations at about 7 and 14 microns from the inner wall of Schlemm's canal the pressure measured at 7 microns was 16.9 +/- 1.3 and at 14 microns it was 18.9 +/- 1.4 cmH2O--that is, 5.2 +/- 0.8 and 7.2 +/- 1.0 cmH2O respectively, higher than the PSc. The results indicate that at the spontaneous IOP about 75% of the resistance between the anterior chamber and Schlemm's canal is located within 14 microns from the canal with some 50% being located within the region 7 and 14 microns from the canal. After a small increase in IOP, the tissue causing most of the outflow resistance became relocated to a region within 7 microns from the canal.


Identification of a 9 kDa gamma-crystallin fragment in human lenses. The degraded polypeptides (M(r) less than 14 kDa) were isolated by a preparative SDS-PAGE method from water soluble (WS) and water insoluble (WI) proteins of human lenses from donors of ages between 5 and 75 years. SDS-PAGE analysis showed the presence of a major 9 kDa polypeptide species that showed an age-related increase in levels in WS-polypeptide preparations. In order to identify the parent crystallin of the 9 kDa polypeptide, the immunoreactivities of the WS- and WI-degraded polypeptides to immuno-affinity-purified anti-human alpha-, beta- and gamma-crystallin antibodies were determined by the Western blot method. The WS- and WI-9 kDa polypeptides showed immunoreactivity to only the anti-gamma-crystallin antibody suggesting it to be a fragment of gamma-crystallin. A 9 kDa species was purified by Sephadex G-50 chromatography from the WS-protein fraction of lenses from 20-30-year-old donors. The purified polypeptide showed a single protein band during SDS-PAGE and also an apparent single spot on two-dimensional gel-electrophoresis (IEF followed by SDS-PAGE). The purified preparation also showed a single major peak during reverse phase HPLC chromatography. The purified 9 kDa polypeptide showed immunoreactivity to only the anti-gamma-crystallin antibody. A polyclonal antibody raised against the purified 9 kDa polypeptide showed immunoreactivity only to a 20 kDa gamma-crystallin species. The partial N-terminal sequence analysis of the 9 kDa polypeptide showed it to be a fragment of gamma D-crystallin. Together these results show that a 9 kDa gamma D-crystallin fragment exists in increasing quantities in human lenses during aging.


Effect of enzymatic deglycosylation on the regenerability of bovine rhodopsin. The influence of the carbohydrate groups of rhodopsin on its ability to regenerate upon incubation with 11-cis retinaldehyde after photobleaching was examined. Rhodopsin was deglycosylated enzymatically with peptide-N-glycosidase F (PNGase F). Verification of deglycosylation was established by: (a) SDS-PAGE; (b) carbohydrate compositional analysis using high performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD); (c) isolation and carbohydrate analysis by HPAEC-PAD and fast atom bombardment-mass spectrometry of the oligosaccharides liberated from rhodopsin; and (d) absence of reactivity with lectins. Deglycosylated rhodopsin, when present either in rod outer segments or after purification, exhibited the same absorption spectrum as the native molecule. After photobleaching, deglycosylated rhodopsin reacted with 11-cis retinaldehyde in a manner similar to the native material, restoring the spectral properties lost after light-exposure. The carbohydrate portion, therefore, was not required for expressing the spectral properties of rhodopsin nor for regeneration of the photobleached visual pigment.


Uptake of 22-carbon fatty acids into rat retina and brain. Rat retina accumulates high levels of 22-carbon (C22) polyunsaturated fatty acids (PUFA), especially docosahexaenoic acid (DHA, 22:6 omega 3), in rod outer segment (ROS) phospholipids (PL). However, plasma, the source of retina lipids, is enriched in 20-carbon (C20) fatty acids instead of C22 PUFA. This suggests that the retina has a mechanism(s) for selective uptake of C22 PUFA from the blood. It is not known if the selective uptake is specific for the carbon number alone, or if the number of double bonds is also important. To address this question, the following study was carried out using erucic acid (22:1 omega 9) as a metabolic marker molecule. Albino rats were raised from birth on a diet containing 10% (by weight) of either rapeseed oil (43% 22:1 omega 9) or blended canola oil (0.4% 22:1 omega 9). At 4 months of age, plasma, liver, adrenal gland, brain and retina were collected, lipids were extracted, and fatty acids were determined. In those rats fed rapeseed oil, 22:1 omega 9 was incorporated into the lipids of plasma (2.3%), liver (0.6%), and adrenal gland (17.6%), indicating that this fatty acid was absorbed, transported, and metabolized by the rats. However, 22:1 omega 9 was not incorporated into the lipids of retinal ROS or brain. Our results suggest that both the carbon number and degree of unsaturation are important determinants in the selective uptake of C22 fatty acids from plasma into both the brain and the retina.


Evidence for a Na(+)-Cl(-)-H(+)-HCO3- exchange system in the mammalian lens. 36Cl- efflux was studied in the isolated rat lens under two conditions that are known to decrease internal pH. The first follows exposure to a pulse of ammonium chloride (50 mM) and the second accompanies exposure to an acidified propionate (20 mM) solution. Under acidifying conditions, a stimulation in 36Cl- efflux was observed, that was abolished on removing external Na+ and also on removing external Cl- and HCO3-. In the absence of external Cl-, the presence of HCO3- (16 mM) resulted in an increase in 36Cl- efflux during internal acidification. In the absence of internal acidification, the addition of 0.1 mM dibutyrylcAMP or 0.5 mM IBMX to the external medium produced a rapid increase in 36Cl- efflux. This stimulation was reduced by 0.2 mM SITS. Neither cAMP or IBMX had any significant effect on the electrical resistance of the lens membranes. It is suggested that a coupled SITS-sensitive, Na(+)-Cl(-)-H(+)-HCO3- exchange mechanism is activated when the lens internal pH falls and further that cAMP may play a role in regulating this mechanism.


Rhodopsin levels in the central retinas of normal miniature poodles and those with progressive rod-cone degeneration. Visual pigment in normal miniature poodles and those with progressive rod-cone degeneration (prcd), a late-onset autosomal recessive photoreceptor degeneration, has been studied using imaging fundus reflectometry (IFR). The stage to which the disease had advanced in the animals with prcd was assessed with electroretinography (ERG). Measurements were carried out on seven affected, two heterozygous and three homozygous normal animals. The IFR measurements showed that the in situ difference spectrum of visual pigment measured in the central retina of the normal poodle is typical of vertebrate rhodopsin, with a maximum at about 510 nm. Rhodopsin regeneration following extensive bleaches continues for up to 70 min. In poodles with prcd, rhodopsin is spectrally normal and regenerates at normal rates. In young affected animals under 1 year of age, the final levels of rhodopsin could already be substantially reduced. Serial measurements of visual pigment in these dogs showed differences in the degree and spatial pattern of pigment loss and rate of progression between animals. The extent of visual pigment loss also differed among the older (greater than 4.5 years) affected animals: while in one animal no pigment could be detected, in another a central band of retina was relatively spared, and significant levels of visual pigment were measured within it. Pigment levels measured within the central 25 degrees of the retinas of poodles heterozygous for prcd were lower than those in normal animals, even though their ERGs were within the normal range.


Synthesis and secretion of interphotoreceptor retinoid-binding protein (IRBP) and developmental expression of IRBP mRNA in normal and rd mouse retinas. The synthesis and secretion of interphotoreceptor retinoid-binding protein (IRBP) was quantitatively assessed in retinas of normal and rd mutant mice using short-term organ culture with [35S]methionine. Retinas were studied at ages P9-P12, time points prior to and immediately after the onset of the degeneration of the rd retina. Soluble proteins of the retinal pellet and the incubation medium were subjected to SDS-polyacrylamide gel electrophoresis. Analysis of labeled protein bands utilized a radioactivity scanning system to quantify [35S]methionine incorporation into newly synthesized IRBP. The synthesis and secretion into the incubation medium of IRBP by rd mouse retinas was comparable to normal retinas at P9-P10 but decreased by more than 50% by P12. IRBP mRNA levels were evaluated in retinas of normal and rd mice ages P7-P14. Although IRBP mRNA expression increased in the rd mouse through P10, it decreased markedly thereafter. Previously reported immunocytochemical studies suggested that IRBP was not secreted in the rd mouse retina. The results of this study indicate, however, that rd mouse retinas, when removed from the eye, have the capacity to synthesize and secrete IRBP.


Morphologic and clinical study of the retinal circulation in the miniature pig. A: Morphology of the retinal microvasculature. The microvasculature of the retina was studied in 20 miniature pigs by means of vascular corrosion casts and semithin histological sections. These techniques provided information useful for a better understanding of retinal fluoangiography. Various characteristics of the porcine retinal vessels, which are similar in the human retinal vasculature, were demonstrated in this study: the holangiotic retina, the trilaminar organization of the retinal capillaries and the presence of particular avascular zones, radial peridiscal capillaries and initial annular constrictions of certain arteriolar side-branches. The intrinsic retinal vasculature also shows some dissimilarities between both species. The major retinal blood vessels of the pig lie very superficially in the nerve fibre layer, in contrast to their deeper location in man. The streak-shaped macular area of the pig contains no major vessels but, unlike the central fovea in the human eye, it is not completely avascular. Another interesting finding is the presence of a large anterior border venule in the porcine retina.


Morphologic and clinical study of the retinal circulation in the miniature pig. B: Fluorescein angiography of the retina. The retinal blood circulation was examined in 20 porcine eyes by studying fluoangiograms before and after occlusion of the temporal choroidal circulation. The use of a small intravenous bolus of 2.5 ml fluorescein provided high-resolution angiograms with detailed information of the retinal vascular bed. The lobular filling pattern of the choriocapillaris very closely resembles the human pattern. The intrinsic retinal vasculature of the miniature pig also has numerous characteristics in common with the human retina with regard to the extent of the vascular bed, the size of the blood vessels and the presence of radial peridiscal capillaries. Unlike the central fovea in the human eye, the porcine retina has a streak-like macular area which extends both temporally and nasally and which, although it is free of major blood vessels, is not completely avascular. This fluoangiographic study substantiates the results of the accompanying paper. It may be concluded that the miniature pig is a suitable experimental animal for applied research on the retinal vasculature.


Recombinant human basic fibroblast growth factor (Rh-bFGF) in three different wound models in rabbits: corneal wound healing effect and pharmacology. Prior to a clinical trial in humans, we studied the effect and pharmacological distribution of recombinant human basic fibroblast growth-factor (Rh-bFGF) in vivo. Healing experiments on de-epithelialized rabbits corneas (n = 24 animals) compared the efficacy of three bFGF doses to controls and revealed a significantly increased healing rate for both 200 ng and 500 ng per application Rh-bFGF treatment groups compared to the control groups. To assess possible side effects of Rh-bFGF (500 ng topically applied for up to 7 days, twice daily), ten rabbits were involved in a model of an anterior keratectomy wound (performed with Draeger's roto-keratome to a depth of 0.15 mm). Light microscopy of thin sections of treated corneas showed an increased fibrogenesis in the anterior stroma with a more pronounced activation of keratocytes. No evidence for abnormal neovascularization or inflammation was observed when compared to control corneas. Ocular penetration and systemic distribution of topically applied labelled 125I FGF was assessed in three models (iodine vapour epithelial burn, anterior keratectomy and penetrating autokeratoplasty) in 24 rabbits. No intraocular penetration of bFGF occurred as shown by direct gamma counting. Macroautoradiography showed a selective labelling of epithelial basement membrane when denuded and intact, as previously described. Evidence for systemic absorption of breakdown products was confirmed by heparin-sepharose chromatography of blood and urine samples. Under these conditions, we suggest that topical Rh-bFGF promotes corneal wound healing without morphological adverse reaction or intraocular and systemic penetration.


Electrical fields in the vicinity of epithelial wounds in the isolated bovine eye. A lateral potential drop along the outer surface of the cornea could be measured at the edge of wounds made in the corneal epithelium of the isolated bovine eye when the cornea was covered by simulated tear film of modified Hanks' solution. These lateral fields (LFs) had an average magnitude of 42 +/- 1.4 mV mm-1 in the first 0.25 mm from the wound edge. The polarity of these surface LFs is more positive at the wound than in regions away from the wound. Very little if any lateral field could be measured at the edge of wounds along the inner surface of the corneal epithelium. The surface LFs depend on the cornea's transepithelial potential (TEP), which in this isolated bovine eye preparation we determined (with conventional microelectrode techniques) to have an average value of 24.7 +/- 2.2 mV, stroma-side positive. We found that this TEP drives a current from wounds in the epithelium with an average current density of 55 +/- 12 microA cm-2. We also have found that the average transcorneal potential (TCP) was 3.2 +/- 0.5 mV greater than the average TEP, confirming that the major contribution to the TCP comes from the corneal epithelium.


Genetic variability and rodent models of human aging. Inbred strains, outbred strains, and natural populations of rodents differ greatly in the amount and nature of the genetic variability they possess. Consequently, as models of human aging they vary with respect to the areas of research to which they are best suited. Inbred strains, in which all individuals are genetically identical, are best suited as models of specific disease processes and for manipulations involving tissue transplantation. Their lack of genetic variability, however, and the disruption of genetic linkage groups that occurs during inbreeding limit their value as models of more general aging processes. Outbred strains exhibit large interindividual genetic variation--a result of ongoing random accumulation of deleterious alleles with late ages of action. This makes them ideal models for studying the diversity of pathologic lesions, connections between pathologies, and susceptibility to pathologic lesions that collectively produce the reductions in reproductive capacity, physiological efficiency, and viability that are characteristic of aging. Natural populations also may exhibit relatively large amounts of interindividual genetic variability. However, difficulties with husbandry, variable parasite loads, and complex population genetics can compromise their suitability as models of human aging. Ultimately, a consideration of the range of animal models available and a more careful matching of the goals of a study with the genetic system of the model will prove fruitful to gerontology.


The reductive capacity index of saliva obtained from donors of various ages. The reducing ability of saliva is decreased with donor age as indicated by cyclic voltammetry measurements of saliva. It has been shown that saliva possesses a strong reducing power that is influenced by the induction of oxidative stress. The exact capacity of saliva to cope with oxidative stress has never been studied in detail. We used a new approach to detect the role of oxidative stress on saliva. This approach included the detection of the total reducing capacity of the saliva without determination of the exact concentration of a specific reducing compound present in the saliva. Induction of oxidative stress resulted in a significant change in the reductive capacity detected. Measurements of the reducing potential of saliva revealed that there is a correlation between the donor age and the reductive capacity observed. Also, saliva obtained from donors in various disease states have shown a significant change in the potential recorded, as compared to controls. The possible use of this method as a diagnostic tool in various diseases is discussed.


Effect of cold on serum substrate and glycogen concentration in young and old Fischer 344 rats. This investigation evaluated the hypothesis that the age-related decline in cold-induced thermogenesis observed in male (F344) rats is associated with altered substrate concentrations of glucose, lactate, and/or liver and muscle glycogen. Body mass-independent O2 consumption, core temperature, and serum glucose and lactate concentrations were measured at rest and during 4 h of exposure to 5 degrees C in male F344 rats ages 6, 12, and 26 months. At the end of the 4-h cold exposure, liver, soleus, and gastrocnemius tissues were removed, frozen, and analyzed for glycogen concentration and/or citrate synthase activity. Core temperature decreased during cold exposure and was consistently less in the 26-month versus the 6- and 12-month rats. There were no significant differences between the 6- and 12-month-old rats with respect to cold-induced O2 consumption, but measures were significantly lower in the 26-month-old rats. During cold exposure, serum lactate and glucose concentrations increased in the 26-month-old animals compared to those in the 6- and 12-month-old rats, while liver glycogen concentrations decreased in all groups, and gastrocnemius glycogen contents decreased in the 12- and 26-month-old rats. Citrate synthase specific activity (mumol.[min.microgram.protein] -1) did not differ with age. These data suggest that carbohydrate availability (as measured by serum glucose and muscle glycogen) is not a limiting factor in the attenuated cold-exposed thermogenic response of the 26-month-old male F344 rat. However, it appears that the 26-month-old rat may have a diminished capacity to fully oxidize carbohydrate during cold exposure.


Lymphoproliferative responses in diet-restricted and aging Sprague-Dawley rats. Immunological effects of aging and dietary restriction (i.e., chronic underfeeding without malnutrition) were investigated in male CFY Sprague-Dawley rats. From weaning, diet-restricted animals were given the amount of food that maintained their body weights at approximately 50% of age-matched ad libitum controls. Cells from the spleen, Peyer's patches, brachial/axillary lymph nodes, and mesenteric nodes of 12- and 20-month-old rats were tested in vitro against the T-cell mitogens phytohaemagglutinin and concanavalin A, and the B-cell mitogen Salmonella minnesota lipopolysaccharide (with dextran sulphate). No clear boosting effect of dietary restriction nor age-related decline in T- or B-cell response was observed when a standard foetal bovine serum supplement was used. In general however, serum from diet-restricted rats supported better proliferative responses than serum from age-matched controls, suggesting that dietary restriction may promote lymphocyte proliferation by an indirect mechanism. A feature of this study was the variation in phenotype and immune responsiveness among closely related animals. Thus in an outbred population, any beneficial effects of dietary restriction upon immune responses could be outweighed by variation among individuals.


Changes in conjugative enzyme activity and acetaminophen metabolism in young and senescent male F-344 rats following prolonged exposure to buthionine sulfoximine. This study examined how advanced age affects glucuronide and sulfate conjugation of acetaminophen after prolonged exposure to L-buthionine-S,R-sulfoximine (BSO) in male Fischer 344 rats. Young (4-5 month) and senescent (21-22 month) rats received 11 doses of BSO (2 mmol/kg) at 12-h intervals via a gastric cannula. Hepatic metabolism was assessed in vivo by measuring the products of reactions mainly responsible for acetaminophen elimination, namely the formation of the glucuronide and sulfate conjugates. Selected drug-metabolizing enzyme activities were also determined in vitro. BSO treatment increased the partial clearance to acetaminophen glucuronide by 90% and 41% in young and old rats, respectively, and similarly, induced p-nitrophenol and 1-naphthol UDP-glucuronosyl transferase activities to a greater extent in young versus senescent animals. Thus, the induction of these UDP-glucuronosyl transferase activities by BSO is preserved in senescent animals. Although the partial clearance to acetaminophen sulfate was decreased in senescent control rats compared to young controls, BSO treatment decreased the in vivo rate of sulfation in both age groups. Similar to previous findings with the Sprague-Dawley strain, BSO treatment did not induce hepatic cytochrome P-450 content or activity or cytosolic p-nitrophenol sulfotransferase activity in young and senescent Fischer 344 rats.


Learning performances and aging in cuttlefish (Sepia officinalis). The use of an associative learning protocol with negative reinforcement allowed us to compare the performances of three groups of animals: young, middle-aged, and senescent. Statistical analysis of the results does not enable us to assert that a significant difference exists between the learning speeds of young and senescent animals. On the other hand however, the latter show poor scores in a retention test carried out 24 h after the learning session. These results suggest that the long-term memory processes are affected during the aging of these animals.


Selection on stress resistance increases longevity in Drosophila melanogaster. Tests for the causal involvement of specific physiological mechanisms in the control of aging require evidence that these mechanisms can be used to increase longevity or reproductive lifespan. Selection for later reproduction in Drosophila has been shown to lead to increased longevity, as well as increased resistance to starvation and desiccation stresses. Selection for increased resistance to starvation and desiccation in Drosophila melanogaster is here shown to lead to increased longevity, indicating that alleles that increase stress resistance also may increase longevity. The responses of desiccation and starvation resistance to selection are partly independent of each other, indicating a multiplicity of physiological mechanisms involved in selectively postponed aging, and thus aging in general.


Increase of basic fibroblast growth factor immunoreactivity and its mRNA level in rat brain following transient forebrain ischemia. We examined the time course of basic fibroblast growth factor (bFGF) immunoreactivity and its mRNA level mainly in the hippocampus after transient forebrain ischemia using immunohistochemistry, enzyme immunoassay (EIA), Western blot analysis and in situ hybridization. Neuronal death in the hippocampal CA1 subfield was observed 72 h after 20 min of ischemia. The number of bFGF-immunoreactive(IR) cells increased 48 h-5 days after ischemia in all hippocampal regions. At 10 and 30 days, the bFGF-IR cells in the CA1 subfield had further increased in numbers and altered their morphology, enlarging and turning into typical reactive astrocytes with the advancing neuronal death in that area. In contrast, the number of bFGF-IR cells in other hippocampal regions had decreased 30 days after ischemia. The EIA study showed a drastic increase in bFGF levels in the hippocampus 48 h after ischemia (150% of that in normal rat) which was followed by further increases. In Western blot analysis, three immunoreactive bands whose molecular weights correspond to 18, 22 and 24 kDa were observed in normal rat and ischemia increased all their immunoreactivities. In the in situ hybridization study of the hippocampus, bFGF mRNA positive cells were observed in the CA1 subfield in which many bFGF-IR cells existed after ischemia. These data demonstrate that transient forebrain ischemia leads to an early and strong induction of bFGF synthesis in astrocytes, suggesting that the role of bFGF is related to the function of the reactive astrocytes which appear following brain injury.


Differential cellular distribution of two sulphur-containing amino acids in rat cerebellum. An immunocytochemical investigation using antisera to taurine and homocysteic acid. An antiserum to homocysteic acid was raised in rabbits. Immunogens were prepared by coupling this amino acid to bovine serum albumin by means of glutaraldehyde and paraformaldehyde. When applied to semithin or ultrathin sections of rat cerebellum, the antiserum produced selective labelling of glial cells and processes, including the Bergmann fibers. No enrichment of immunoreactivity was detected in nerve terminals of the major excitatory fiber systems. The distribution of homocysteic acid-like immunoreactivity was very different from that of taurine (another sulphur-containing amino acid), as judged from consecutive semithin sections labelled with a postembedding immunoperoxidase procedure and from ultrathin sections labelled with a postembedding double immunogold procedure. Taurine-like immunoreactivity was concentrated in Purkinje cells and was low in glial elements. Our data suggest that the cerebellum contains a glial pool of homocysteic acid (and/or precursors that may undergo spontaneous oxidation to homocysteic acid) and that this amino acid is unlikely to act as a cerebellar transmitter.


Plasticity of binocular visual connections in the frog, Xenopus laevis: reversibility of effects of early visual deprivation. In frogs, each lobe of the midbrain optic tectum receives two spatially aligned maps of binocular visual space, one through each eye. The map from the contralateral eye is the product of direct retinal input to the tectum, while that from the ipsilateral eye is mediated by a commissural system of connections that relays binocular visual information from one tectal lobe to the other. In Xenopus laevis, dark-reared from early life, the visuotectal map through the ipsilateral eye exhibits significant signs of disorder indicating that the normal development of the underlying "intertectal" system involves experience-dependent processes. Here we investigate the potential of this system for recovery from such deprivation-induced disruption. Three groups of animals were first dark-reared from embryonic stages until 3 months, 12 months or 2 years after metamorphosis. These short-, medium- and long-term periods of visual deprivation are associated with a progressive disturbance of the intertectal system (Grant and Keating 1989b). The animals then experienced 12-18 months in a normal visual environment, before an electrophysiological mapping experiment was carried out to reveal the status of their intertectal system. Quantitative analyses of the visuotopic order in the ipsilateral visuotectal map and of its spatial alignment with the map derived from the contralateral eye revealed that these features were completely normal in all 3 experimental groups. It is concluded that defects present in the intertectal system of dark-reared Xenopus can be repaired by subsequent visual experience, and that the reparative capacity is not limited either by the animal's age or by the existing degree of prior disruption.


Binocular interaction and disparity coding at the 17-18 border: contribution of the corpus callosum. Binocular disparity, resulting from the projection of a three-dimensional object on the two spatially separated retinae, constitutes one of the fundamental cues for stereoscopic perception. The binocularity of cells in one hemisphere stems from two sources: i) from the ipsilateral ganglion cells in the temporal retina which converge with inputs coming from the contralateral nasal retina; the latter axons cross at the chiasma; ii) from inputs originating in the opposite hemisphere which cross in the corpus callosum. The objective of this study was to demonstrate that interactions from both types of inputs can result in the formation of disparity sensitive neurons and presumably that either type could mediate stereoperception based on disparity cues. Two types of disparity sensitive neurons were found in the normal cat: one type, showing maximal interactive effects around zero disparity responded with strong excitation or inhibition when the stimuli were in register. These neurons are presumed to signal stimuli situated about the fixation plane. The other type, also made up of two subtypes of opposed valencies, gave maximum responses at one set of disparities and inhibitory responses to the other set. These were presumed to signal stimuli situated in front of or behind the fixation plane. In the split-chiasm cat, whose cortical binocularity is presumably assured by converging ipsilateral and callosal inputs, three of the four subtypes of disparity sensitive neurons were found, the uncrossed disparity cells being absent in these animals. Moreover, stimulating each eye individually indicated that nearly 80% of the cells in normal and about 40% in split-chiasm cats were binocularly driven. However, both these figures underestimated the amount of binocular interaction in the callosal recipient zone, since stimulating both eyes simultaneously showed that a proportionately larger number of cells were binocularly driven. Disparity sensitive cells also varied as a function of ocular dominance, i.e., cells signaling the fixation plane tended to have balanced dominance whereas units preferring stimuli situated in front of or behind the fixation plane were dominated by the ipsilateral and contralateral eyes, respectively.


An improved graphical method for pattern recognition from spike trains of spontaneously active neurons. Spontaneous activity and rhythmical oscillations are common features of large neuronal networks in mammals. Detection of repetitive spike patterns or pacemaker activity during electrophysiological recording of spontaneous action potentials from single neurons can be difficult if a "noisy" background is present. This paper describes an improved method for an online spike train analysis based on joint interval histograms (JIH, Rodiek et al. 1962). By means of higher ordered JIH the discrimination of spike patterns with repetitive bursting activity or oscillations is possible even when randomly distributed action potentials appear. Examples of simulated spike trains and those recorded from cultured hippocampal neurons are presented.


Midbrain influences on ventrolateral medullo-spinal neurones in the rat. A study has been carried out to investigate the influence of the periaqueductal grey matter (PAG) on spinally-projecting neurones in the rostral ventrolateral medulla (RVLM) in rats anaesthetised with urethane. Microinjection of D,L-homocysteic acid was used to selectively activate nerve cell bodies in the PAG. Stimulation throughout the rostral half of the PAG and in the caudal PAG on a level with and dorsolateral to the level of the aqueduct evoked excitatory responses in 21 medullo-spinal cells in the RVLM. The neuronal excitation was associated with a rise in blood pressure. In contrast, stimulation within a restricted area of the ventrolateral PAG at the level of the dorsal raphe nucleus inhibited 10 medullo-spinal neurones in the RVLM, and produced variable changes in blood pressure. Convergence of excitatory and inhibitory influences from dorsolateral and ventrolateral stimulating sites in the PAG on to individual cells was also demonstrated. The results are discussed with respect to the role of the ventrolateral PAG in modulating the cardiovascular components of the "defence" response which is integrated by the dorsal PAG.


Role of upper cervical inspiratory neurons studied by cross-correlation in the cat. Axonal projections and synaptic connectivity of upper cervical inspiratory neurons (UCINs) were investigated in anaesthetised cats to clarify their role as propriospinal respiratory interneurons. Antidromic mapping showed axonal collaterals near phrenic and intercostal motonuclei. Of the UCINs tested, 37% had collaterals at T3-4; 55% had ipsilateral projections and 45% had contralateral projections. Ipsilateral or contralateral cross-correlations of the activity of pairs of UCINs (one on each side of the spinal cord) with the discharge of internal intercostal, external intercostal (T3-4) or phrenic nerves revealed similar features. Those with the internal intercostal and phrenic nerves were interpreted as evidence for shared or oligosynaptic excitation, those with the external intercostal nerve as shared excitation and inhibition. No evidence for monosynaptic connections was found. Monosynaptic connections could also not be demonstrated between inspiratory intercostal neurons located near (less than 0.5 mm) the UCINs collateral arborizations in T3-4, examined by cross-correlation. Afferent feedback from internal intercostal nerves (T3-4) was investigated by cross-correlating nerve stimulation with UCINs activity. Ipsilateral and contralateral cross-correlograms had similar features, providing evidence for excitation in some cases and inhibition in others. Finally, cross-correlations between ipsilateral UCINs and cervical sympathetic nerves were featureless. The results suggest that the role of UCINs as part of a respiratory propriospinal control system analogous to forelimb motor control is untenable, although they may be part of an intercostal afferent feedback loop.


Nucleus basalis lesions in neonate rats induce a selective cortical cholinergic hypofunction and cognitive deficits during adulthood. Ibotenic acid was infused into the nucleus basalis magnocellularis (nBM) of 2-day old rats to eliminate immature cholinergic neurons before they develop functional synaptic connections in the neocortex. For bilaterally lesioned neonates, cognitive testing was initiated 2 months after lesioning and animals were sacrificed at 8 or 12 months of age. Lesioned animals exhibited a marked deficit in the retention of passive avoidance behavior, as well as in the acquisition of 2-way active avoidance behavior. Lesioned animals also made significantly more alternation errors than control animals in the Lashley III spatial maze and showed severe impairments in general learning, reference memory and working memory during 17-arm radial maze testing. For all 4 tasks, neonatally lesioned animals did not show any recovery to the performance level of control animals. Histological analysis of the subcortex from lesioned animals during adulthood revealed: (1) a substantial reduction in acetylcholinesterase-positive cells (presumably cholinergic) within the nucleus basalis, (2) decreased acetylcholinesterase staining in neocortex and (3) a gliosis essentially restricted to the globus pallidus. Surrounding brain regions were apparently not damaged as a direct result of excitotoxin infusion. Neurochemically, neonate nBM lesioning produced a long term cholinergic hypofunction as evidenced by significant reductions of 25% and 18% in frontal cortex choline acetyltransferase (CAT) activity at 12 and 8 months of age, respectively. By contrast, prefrontal cortical concentrations of biogenic amines and their metabolites were unaffected, thus indicating a degree of neurochemical specificity for these neonatal nBM lesions. The persistent cortical cholinergic hypofunction in lesioned animals may be related to the long term deficits in learning/memory abilities that were also observed. It is suggested that neonatal nBM lesioning could provide a useful animal model for elucidating the plasticity of the developing brain after cortical anervation.


Human ocular torsion during parabolic flights: an analysis with scleral search coil. Rotation of the eyes about the visual axis is known as ocular torsion. A lateral inclination (a "roll") of the head induces ocular torsion in the opposite direction, a response known as ocular counterrolling. For six subjects, we recorded the static (head still) and dynamic (head in oscillatory roll motion) ocular torsion in normal 1 g condition and also during the microgravity and hypergravity periods of parabolic flight, using the electromagnetic scleral search coil technique. With the head still, the direction and magnitude of torsion that occurred in response to microgravity and hypergravity differed substantially from one individual to another, but there was a significant difference in torsional magnitude between the microgravity and hypergravity periods, for all static head positions including the upright position. Under normal 1 g conditions, counterrolling compensated for about 16% of (voluntary) static head roll, while dynamic counterroll was much larger, up to 36% of head roll at 0.55 Hz. With increasing frequency of head oscillation between 0.33 Hz and 0.55 Hz, the gain of counterrolling increased and there was no change in the phase relationship. The gain of dynamic counterroll (in response to voluntary head rolling) was not significantly less in hypogravity, suggesting that on the ground at these frequencies the contribution of gravity and gravity receptors to this reflex is redundant: this reflex is probably driven by the semicircular canals. In some subjects, the torsional displacement in microgravity is accompanied by micro-torsional oscillatory motion.


Effect of knee joint laxity on long-loop postural reflexes: evidence for a human capsular-hamstring reflex. The onset latency and discharge amplitude of preprogrammed postural responses were evaluated in order to determine if the structure of synergistic activation could be altered by ligamentous laxity at the knee joint. Twelve subjects with unilateral and one subject with bilateral anterior cruciate ligament (ACL) insufficiency were tested while standing on a moveable platform. External balance perturbations (6 cm anterior or posterior horizontal displacements of the platform) were presented at velocities ranging from 15 to 35 cm/s. Perturbations were presented under the following experimental conditions: unilateral and bilateral stance, knees fully straight or flexed, and with ankle motion restricted or free. These stance, knee position, and ankle motion conditions were introduced to alter the stress transmitted to the knee joint during movement of the support surface. The automatic postural response was recorded from the tibialis anterior (T), quadriceps (Q), and medial hamstrings muscles (H) bilaterally. The normal response to an externally induced backward sway involved the automatic activation of T and Q at latencies of 80 ms and 90 ms respectively. Activation of the hamstrings in the non-injured extremity was not coupled with the postural response. Hamstrings are not typically involved in the correction posterior sway because H activation would tend to pull the center of mass further backwards. However, when the response in the ACL-deficient extremity was compared to the non-injured limb: (1) the automatic postural response in the ACL-deficient extremity was restructured to include hamstrings activation (100 ms latency), (2) H activation time was faster and less variable in the ACL-deficient limb, and (3) the ratio of H/Q discharge amplitude integrated over 100 ms and 200 ms from the onset of EMG activation showed a dominance of hamstring activity during unilateral stance on the lax limb. In addition, H/Q ratios integrated over 200 ms showed dominant hamstring activity in the ACL-deficient limb during bilateral stance. (4) Cross-limb comparisons showed greater normalized IEMG amplitudes for T, H, and Q during unilateral stance on the lax limb. These results suggest that a capsular-hamstring reflex is integrated into the existing structure of a preprogrammed postural synergy in order to compensate for ligamentous laxity. Furthermore, the generalized increase of response gain observed during perturbations of unilateral stance on the lax limb indicates that joint afference can modulate central programming to control localized joint hypermobility. A concept of postural control is discussed with respect to the capsular reflex, joint loading and displacement of the center of gravity.


Inhibitory effects on flexor reflexes in patients with a complete spinal cord lesion. The inhibitory effects on flexors of electrical stimulation of a distal peripheral nerve were investigated in 7 paraplegic patients having a complete spinal cord section. The stimuli (3-50 mA) were applied to the sural nerve. Their effects were investigated on: 1) the ipsi- and contralateral H reflex of the Tibialis Anterior (TA); 2) the continuous EMG activity reflexly elicited in TA by a sustained pinch of the foot and 3) on the reflexes evoked in TA by contralateral sural nerve stimulation. Sural nerve stimulation induced two peaks of facilitation of the ipsilateral TA H reflex that could be replaced by inhibition as the stimulus intensity was increased. The comparison of the effect on H reflexes and the EMG activity suggests presynaptic inhibition of Ia fibres at time intervals longer than 300 ms. The stimulation could depress the sustained EMG reflex activity and induce a period of silence whose duration increased with the intensity of the stimulation. As shown in a previous study, a sural nerve stimulation induced a reflex in TA with a prolonged (more than 130 ms) latency. This late reflex could be selectively inhibited by a contralateral sural nerve stimulation, probably at an interneuronal level. These results confirm that the late reflex in TA is similar to the one observed after Flexor Reflex Afferent (FRA) stimulation in the acute spinal cat with DOPA. In addition, they show that at least some part of the "half centre" organization which has been described in the acute spinal cat with DOPA is also present in the human spinal cord chronically deprived of supraspinal control.


Motor control prior to movement onset: preparatory mechanisms for pointing at visual targets. The present study investigated the mechanisms involved in the preparation of pointing movements in humans. We provided visual precues on the location of the upcoming target, and registered the effect of these precues on the reaction time (RT = interval between target appearance and movement onset). Generally, precues were found to reduce RT, suggesting that some aspects of the preparatory process have been advanced in time. In Exp. 1, precues fully specified the direction required for the upcoming movement while indicating only a range of movement amplitudes; in Exp. 2, precues fully specified the amplitude and indicated a range of directions. In both experiments, RT was shorter than in control trials without precues, and gradually increased with the size of the precued amplitude or direction range. This result suggests that the preparation of either parameter is possible without knowing the precise value of the other, i.e. amplitude and direction are not prepared in a fixed order. Furthermore, our results are consistent with the view that movement preparation includes a progressive contraction of the precued range towards the final value. The speed of this process can be estimated as 0.31 cm/ms for amplitude, and 1.7 deg/ms for direction ranges. In Exp. 3 and 4, precues indicated both amplitude and direction as ranges only. The size of the amplitude range was held constant while the size of the direction range was varied (Exp. 3), or vice versa (Exp. 4). Under these conditions, RT increased with the size of the varied range.(ABSTRACT TRUNCATED AT 250 WORDS)


Firing relations of medial entorhinal neurons to the hippocampal theta rhythm in urethane anesthetized and walking rats. The firing of neurons from layers II and III of medial entorhinal cortex (MEC) was examined in relation to the hippocampal theta rhythm in urethane anesthetized and walking rats. 1) MEC neurons showed a significant phase relation to the hippocampal theta rhythm in both walking and urethane anesthetized rats, suggesting that this region contributes to the generation of both atropine-resistant and atropine-sensitive theta rhythm components. 2) The proportion of phase-locked cells was three times greater in walking rats (22/23 cells) as compared to anesthetized rats (8/23 cells), indicating that MEC cells made a greater contribution during walking theta rhythm. This difference was also manifest in the greater mean vector length for the group of phase-locked MEC cells during walking: 0.39 +/- 0.13 versus 0.21 +/- 0.08. Firing rate differences between walking and urethane conditions were not significant. 3) In walking rats, MEC cells fired on the positive peak of the dentate theta rhythm (group mean phase = 5 degrees; 0 degrees = positive peak at the hippocampal fissure). This is close to the reported phases for dentate granule and hippocampal pyramidal cells. The distribution of MEC cell phases in urethane anesthetized rats was broader (group mean phase = 90 degrees), consistent with the phase data reported for hippocampal projection cells. These findings suggest that medial entorhinal neurons are the principal determinant of theta-related firing of hippocampal neurons and that their robust rhythmicity in walking as compared to urethane anesthesia accounts for EEG differences across the two conditions.


The effects of wrist muscle vibration on human voluntary elbow flexion-extension movements. The effect of forearm muscle tendon vibration during alternating step flexion-extension movements about the elbow was studied in normal humans. In one experiment, a vibrator was mounted over either the forearm flexor or the extensor muscle. In a second experiment, a vibrator was mounted over either the forearm muscle or the biceps muscle. In both experiments, vibration was applied either to a single muscle or simultaneously to both muscles during elbow flexion-extension movements. After a period of practice, subjects learned the required movements and were able to make them with their eyes closed. Application of vibration to the forearm and the biceps muscle during extension movements produced an undershoot of the required end-movement position. Moreover, application of high-frequency vibration (100 Hz) to the forearm extensor and flexor muscle produced an overshoot of the required end-movement position. The observed results are consistent with vibration induced activation of muscle spindle receptors not only in the lengthening muscle during movement but also in the forearm muscles. It is suggested that the pattern of distribution of muscle spindle afferent from the forearm muscle onto alpha-motoneurons of muscles acting at the elbow has played an important role of alternating step flexion-extension movements.


Memory-guided saccades: what is memorized? In order to find out whether extraretinal (oculomotor, internal) input suffices to provide the oculomotor system with the information necessary for saccadic control, two subjects were asked to make memory-guided saccades in complete darkness, after three different location acquisition conditions. These conditions were visually-guided saccades (SA), providing retinal (external) and extraretinal input, visual peripheral target presentation during central target fixation (FI) (external input only), and smooth pursuit (PU) (internal input only). Either 2 or 12 s (delay) after locating the target, the subjects had to make a memory-guided saccade toward it in complete darkness. The results show that whereas these memory-guided saccades were quite accurate for trials with preceding external input, this was not the case with acquisition through internal input alone. Moreover, the accuracy of memory-guided saccades decreased when the delay increased from 2 to 12 s for both conditions with retinal input, whereas the accuracy increased for the one condition without retinal input, i.e., the smooth pursuit location acquisition. Furthermore, when both retinal and oculomotor inputs were provided, better accuracy of the memory-guided saccades was observed than with single input.


The development of corticospinal projections to tail and hindlimb motoneurons studied in infant macaques using magnetic brain stimulation. The EMG responses evoked in tail, leg and foot muscles by magnetic stimulation (MS) of the brain were investigated in two male macaque monkeys under ketamine sedation. The animals were studied longitudinally over a period of 7 months (from 2.75 to 9.75 months old). MS was generally ineffective in eliciting responses when the animals were 2.75-4.5 months old, even at maximum stimulator output (1.5 Tesla). After this time the threshold for evoking EMG responses decreased considerably, and there was an increase in the probability of occurrence of the responses. These age-related changes plateaued at about 7.5 months, after which they remained fairly constant at adult levels. In both animals the maturation of these responses in tail and hindlimb muscles occurred later than in forelimb muscles.


Control of arm movement after bilateral lesions of area 5 in the monkey (Macaca mulatta). The effect of bilateral area 5 lesions on the analysis of proprioceptive information and the guidance of reaching movements was studied in three rhesus monkeys. In the first paradigm (Proprioceptive discrimination test) the monkeys were trained to discriminate between movements of a joystick to the right or left without visual control; they reported the direction of movement by touching or not touching a screen (go/no-go task). After area 5 had been removed, the monkeys were only mildly impaired on this test. It is concluded that such simple joint movement could be analysed in area 2, area 5 being concerned with more complex arm movements. In the second paradigm (Searching test) the monkey had to find a peanut on a board in the dark using proprioceptive information stored in memory during previous trials. After area 5 lesions, the number of correct reaches was not modified but the number of errors after an incorrect trial (correcting movement) was significantly increased. The data suggests that when visual input is not available, area 5 is involved in the guidance of arm movements on the basis of proprioceptive inputs.


The currents that flow in the somatosensory cortex during the direct cortical response. A current-flow and current-source-density analysis of the sensory evoked response (SER) and the direct cortical response (DCR) in the somatosensory cortex of rats was performed to determine the origin of these potentials. The SER was found to originate in layers II and III, as in cats, with a single excitatory neuronal circuit component. The DCR, on the other hand, has five components, three inhibitory and two excitatory. The activation and magnitude of these components vary with stimulus strength and frequency. During the second and fourth ms of the response, two inhibitory currents flow in layers V and VI; 2 ms later, excitatory current flows in layers II and III. This excitatory current appears to be the same one involved in the SER. Five ms later, the superficial excitatory current is replaced by an inhibitory one in the neighborhood of the DCR's negative peak. At strong stimulus strengths, this is followed by an excitatory current in layer V. The early inhibitory and excitatory components step up through layers upper-VI, V and III over time, implying that inhibition followed by excitation moves upward through cortex. The currents associated with the DCR in somatosensory cortex are compared with those for the DCR in motor and association cortex.


Spatiotemporal characteristics of direction-selective neurons in the middle temporal visual area of the macaque monkeys. In an attempt to elucidate the mechanisms of directional selectivity in the neurons of the middle temporal visual area (MT) of macaque monkeys, we presented small numbers of sequentially flashed stimuli with various temporal and spatial intervals within the receptive field (RF) of direction-selective MT neurons. Experiments were performed using awake macaque monkeys trained to fixate on a set of short stationary lines. Stimuli were presented on a CRT screen under computer control. In two-flash experiments, responses to a test flash presented in the center of the RF were examined following a conditioning flash presented in various locations within the RF. Inhibition in the null direction was observed in about 78% of MT neurons, while facilitation was relatively weak in this group of neurons. In most of these neurons, the ranges of temporal and spatial intervals that produced directional selectivity in two-flash experiments were within half the values and double the values, respectively of those in multi-flash experiments. In the remaining 22% of direction-selective MT neurons, several flashed stimuli were necessary to produce directional selectivity. Most of these neurons showed facilitation in the preferred direction. It appears that the inhibitory mechanisms in the null direction are sufficiently strong to be induced by a single conditioning flash whereas the facilitatory mechanisms are weaker and several stimuli are required for production of the direction-selective response.


The pretectal cholinergic system is involved through two opposite ways in frog monocular OKN asymmetry. Frog monocular horizontal optokinetic nystagmus (OKN) has been studied by coil recordings, before and after unilateral microinjection of cholinergic drugs into the pretectum. The recorded eye was either contralateral or ipsilateral to the injected structure. Before injection, monocular OKN displayed a directional asymmetry, reacting only to stimulations in the temporonasal (T-N) direction. The intrapretectal administration of a cholinergic muscarinic agonist (oxotremorine), as well as that of a nicotinic antagonist (D-tubocurarine), abolished the monocular OKN asymmetry, inducing the appearance of the naso-temporal (N-T) component; the difference between the slow phase velocity gain of both components was no longer significant. These data suggest that acetylcholine (ACh), at the level of the pretectum, acts in opposite ways through muscarinic and nicotinic binding sites; monocular OKN asymmetry could result, at least partially, from a facilitating nicotinic effect and an inhibitory muscarinic effect. Possible interactions with other transmitter systems are discussed.


Orienting behavior in hamsters with lesions of superior colliculus, pretectum, and visual cortex. We examined cortical and subcortical mediation of visual locomotor orienting function by comparing the behavior of hamsters with discrete bilateral lesions affecting the pretectum, superior colliculus (SC), or visual cortex (VC). Orienting and approach to stationary targets was evaluated by measuring the accuracy of hamsters' approaches to small black apertures, located at eye level along the wall of a circular white arena. Hamsters with bilateral ablation of the visual cortex were slightly impaired for approaches to central field targets, whereas those with ibotenic acid lesions of the pretectum (which spares fibers of passage and thus leaves tectal afferents intact) were totally unimpaired. Hamsters with transection of the brachium of SC (BSC) at the prectectal-SC (PT-SC) border were severely impaired in their ability to approach stationary targets in central and peripheral fields. Thus, we did not detect any of the central field sparing that has been reported by others for rodents with similar lesions. Several possible reasons for the disparity between our results and those of others are discussed. Overall, our results indicate that in hamsters the SC is essential for normal visually guided approach to dark, stationary targets throughout the visual field. Further, our results and qualitative observations indicate that the approach errors are most likely due to deficits of visuomotor integration rather than to a lack of visual scanning.


Malignant melanoma of the mucous membranes. Primary malignant melanoma occurring in mucous membranes is uncommon. Of 410 patients with malignant melanoma, 14 (3.4%) had their primary tumour located in mucosal membranes. Five of the tumours were in the mucosal lining of the head and neck, five in the gastrointestinal tract mucosa and four in the female genitalia. Eight of the tumours were ulcerated, necrotic and deeply invasive. The most common histological subtype was acral lentiginous melanoma. Surgical resection of the primary tumour was performed in 10 patients. In the other four, surgery was not feasible, and they were treated by irradiation and immunotherapy. Six patients underwent regional lymph node dissection. In four of them, regional metastases were found. Chemotherapy was given to seven patients for widespread metastases. Nine patients (64%) died of metastatic melanoma within 2 years. The remaining five patients are alive after 3 to 8 years. It appears that mucosal membrane melanoma is a distinct variant of the disease due to both its aggressive biological behaviour and its relative inaccessibility for surgical removal, and may therefore be classified separately.


Aberrant retinal projections to midbrain targets mediate spared visual orienting function in hamsters with neonatal lesions of superior colliculus. Rodents, cats, and most nonmammalian vertebrates with bilateral tectal deafferentation or ablation in adulthood are extremely deficient at orienting to visual stimuli; yet animals with neonatal lesions of superficial layers of the superior colliculus (SC) show partial sparing of this response, particularly for targets in the central visual field. In this study, we sought to determine whether these spared orienting abilities are mediated by aberrant retinal projections to the remaining intermediate layers of the SC, or whether visual cortex (VC) mechanisms or alternative behavioral strategies are responsible. Neonatal golden hamsters received either bilateral heat lesions of the SC (rlSC), or a heat lesion of the right SC and enucleation of the right eye (rSCrE). This latter procedure causes axons from the left eye to recross the tectal midline and terminate in the "wrong" (left) SC (Schneider 1973). As adults, both groups of hamsters were extremely deficient in visually guided approach to stationary targets, although rlSC-lesioned hamsters showed some sparing for central field targets and rSCrE-lesioned hamsters often made wrong-direction turns for targets in the left peripheral field. We then subjected both groups of neonatally lesioned hamsters to bilateral aspiration lesions of the VC. Retesting showed no change in visual orienting behavior as a result of the cortical lesions. Labeling of the optic tract with horseradish peroxidase (HRP) revealed abundant aberrant retinal projections to remaining intermediate layers of the SC and thalamic nucleus lateralis posterior (LP), as well as supernormal innervation of pretectal nuclei, the dorsal terminal nucleus of the accessory optic tract, and the ventral nucleus of the lateral geniculate body (LGv). We conclude that the spared visual orienting capabilities of hamsters with rlSC and rSCrE lesions are mediated by the aberrant midbrain projections, and that cortical mechanisms are not involved in spared visual orienting functions following these neonatal lesions.


Primary non-Hodgkin's lymphoma of the thyroid. Eighty seven cases of primary non-Hodgkin's lymphoma of the thyroid presenting to the Beatson Oncology Centre were reviewed. Stage IE and Stage IIE disease accounted for 79 of 87 cases. In a univariate analysis the presence of dysphagia, dyspnoea, positive nodes, stage or male sex all had a statistically significant detrimental influence on survival. Stage and dysphagia were the most influential individually and a multivariate analysis indicated that the prognostic information in all these features was essentially captured by just these two. A prognostic scoring index based upon stage of disease and the presence of dysphagia has been developed. Overall 5 year survival was 43%. Five year survival for Stage IE patients without symptoms of compression was 74%. Patients who underwent surgical resection of tumour followed by local irradiation appeared to survive longer than patients managed by irradiation without surgery, although after adjustment for prognostic features this advantage was not statistically significant.


Radical breast surgery with a contact Nd:YAG laser scalpel. The use of a contact Nd:YAG laser scalpel during radical and modified radical mastectomy was evaluated in 18 patients with carcinoma of the breast. The laser scalpel performed well as a haemostatic tool, the associated mean blood loss for modified radical mastectomy being 132 ml. Operating time, operative blood loss, laser energy required and postoperative wound drainage were all related to patient obesity, correlating significantly with body weight and/or breast weight. The incidence of axillary seroma was not reduced by laser surgery and occurred in 53% of patients undergoing modified radical mastectomy. The Nd:YAG laser scalpel is an excellent haemostatic tool but it does not appear to have any other advantages over conventional surgery for mastectomy.


Psychological morbidity in the first year after breast surgery. In this prospective study, the psychological morbidity associated with the treatment of breast cancer was assessed. The study population comprised all patients referred to one centre with a recently diagnosed breast lump, who were to undergo surgery. Psychological morbidity was assessed preoperatively and at 6 and 12 months postoperatively by modified Rotterdam Symptom Checklist. Three hundred and twenty patients completed all three questionnaires: 93 women undergoing mastectomy, 73 women having conservation therapy for breast cancer and 156 women having biopsy for benign breast disease. Patients with a breast malignancy smaller than 4 cm in diameter were treated by lumpectomy and radiotherapy, anti-oestrogen therapy or chemotherapy alone or in combination. Psychological morbidity among patients with malignant disease was significantly greater than that seen in the group with benign disease. Among cancer patients, a significant decrease in anxiety and depression occurred during the year following surgery. The study failed to demonstrate any psychological advantage associated with breast conservation.


An analysis of the diagnostic accuracy of endoscopic biopsy and cytology in the detection of oesophageal malignancy. The role of combined cytohistological examination in the differentiation of benign from malignant oesophageal mucosal lesions was studied in 331 patients. Malignancy was confirmed by specimen histopathology in 58 patients. Both endoscopic biopsy and cytology were positive in 41 (71%) patients. Endoscopic biopsy alone suggested malignancy in 10 cases (17%) while cytology was positive after negative biopsy in seven (12%). Cytology yielded four false positive and 10 false negative results giving a sensitivity level of 81%, a specificity of 98% and a positive predictive value for oesophageal malignancy of 92%. Histology on the other hand yielded one false positive and seven false negative results providing a sensitivity of 87%, a specificity of 99% and a positive predictive value of 96%. No patients were left undiagnosed using both diagnostic modalities; cytology increased the diagnostic yield from 87% to 100%. Exfoliative cytology was most valuable in the diagnosis of tumours of the lower one-third of the oesophagus where seven of 31 malignancies (23%) were identified by this method alone (P less than 0.05). Neither the histological type of the tumour nor the morphological appearance was found significantly to affect the diagnostic yield (P greater than 0.05). We conclude that cytological examination should be standard practice in the investigation of oesophageal lesions.


Temporary transection of the trachea in cervical esophagectomy. A new method to preserve the larynx and to achieve a better exposure of the cervical esophagus was performed in four patients with cervical esophageal carcinoma. The trachea, which prevented exposure of the esophagus, was cut, keeping the continuity of the recurrent laryngeal nerves. The tracheo-esophageal space was opened like a double door. The cervical esophagus was replaced with a substitution followed by tracheal reconnection. Three different substitutions were used for the reconstruction of the cervical esophagus: free jejunum, free skin tube graft and stomach. This procedure offers a further excision of the esophagus while preserving the larynx, and results in free surgical margins and safe anastomoses of the alimentary tract. This method is applicable for a curative operation of carcinoma without invasion to the trachea to offer the patient a better quality of life.


Increasing incidence of adenocarcinoma of the proximal stomach. A retrospective study was performed on patients with histologically proven gastric adenocarcinoma during the periods 1972 to 1976 (n = 368) and 1986 to 1990 (n = 870) to analyse the potential changes in tumour distribution within the stomach. A recent increase in gastric cancer patients admitted to our institution represents a change in the referral patterns of the patients. The mean age and male:female ratio according to site distribution of the tumour were not found to differ between the two periods. The proportion of carcinomas of the upper third of the stomach was, however, significantly increased (P less than 10(-6)) in the recent series (36.4%) compared with the old series (20.6%). Since the incidence of gastric cancer in our population seems to be unchanged, this may suggest a true increase in proximal gastric tumours. The data may indicate possible changes in aetiological influences.


The impact of intraoperative ultrasonography of the liver on the surgical strategy of patients with gastrointestinal malignancies and hepatic metastases. Intraoperative ultrasonography (IOUS) of the liver is a very useful tool during surgery for gastrointestinal tumors or hepatic tumors. In a prospective study of 82 elective operations for 57 gastrointestinal tumors and 25 hepatic tumors or metastases, we compared IOUS with preoperative ultrasonography, CT-scanning and MRI. IOUS supplied additional information in 38% of the operations. In 22% of the cases, this had a major impact on the intraoperative strategy with an alteration of the procedure. IOUS has a higher detection rate of intrahepatic lesions, discriminates better between solid or cystic nature of the tumor and offers the possibility of performing a safe and radical hepatic resection by its ability to localize lesions in relation to vascular structures. IOUS is a safe and simple technique which will be more cost-effective than extensive preoperative evaluation of the liver. It provides a more complete clinical staging of patients for whom adjuvant therapy is considered after resection of a colorectal carcinoma.


The effect of portal venous flow on the washout of a regionally injected marker substance 99mTc-methylene diphosphonate after hepatic arterial blockade with degradable starch microspheres. Patients with hepatic metastases derived from colorectal carcinoma have a poor prognosis. Regional chemotherapy, either alone, or combined with agents such as degradable starch microspheres (DSM) that reduce or abolish intrahepatic arterial flow and potentiate the delivery of cytotoxics to hepatic metastases, have not significantly improved survival. We have investigated one positive mechanism, namely the effect of portal venous washout of cytotoxics, for the poor efficacy of drugs administered either alone or in combination with DSM via the hepatic artery in the rat. Using a radiolabelled marker, 99mTc-methylene diphosphonate (MDP), to represent a cytotoxic drug, the initial studies indicated that with the hepatic artery and portal vein clamped, a volume of 0.05 ml of the marker administered via the hepatic artery resulted in the most uniform intrahepatic distribution with minimal washout into the systemic circulation (21 +/- 3.7%). When the hepatic artery was clamped, the washout of MDP was reduced from 100% (with clamps on the portal vein and hepatic artery) to 84.2 +/- 7.7%. DSM administered concomitantly with MDP, resulted in a greater reduction of the portal venous washout of the marker (63 +/- 2.4%). Administration of DSM and MDP via the hepatic artery and with the portal vein clamped further reduced the washout of the marker to (21 +/- 2.26), results similar to those observed with inflow vessel clamps. Following restoration of portal venous flow, there was a rapid washout of 53.7 +/- 7.6% of the marker into the systemic circulation. The results of this study suggest that portal venous washout of regionally delivered cytotoxics, either alone or in combination with DSM, offer an explanation for the poor efficacy of regional chemotherapy in improving the prognosis of patients with hepatic metastases.


Iodostearic acid in the treatment of advanced gastrointestinal carcinoma. This phase II non-comparative trial evaluated the efficacy of intramuscular iodostearic acid in the treatment of 30 patients with advanced carcinoma of the gastrointestinal tract. These included 17 patients with colorectal carcinoma, seven with pancreatic carcinoma, four with gastric carcinoma, one with hepatocellular carcinoma and one with cholangiocarcinoma. Iodostearic acid emulsion (2 ml/1.2 g) followed by depomedrone (0.5 ml/10 mg) was injected intramuscularly twice weekly. No serious toxic effects were observed, the main side effects being local pain at the injection site and hot flushes. There were no partial or complete responses. Stable disease was seen in six of 30 patients (20%) at 3 months and three of 30 patients (10%) at 6 months. In conclusion, intramuscular iodostearic acid is relatively non-toxic, however, it appears to be of little value in the treatment of patients with advanced gastrointestinal carcinomas.


Local excision of rectal cancer: report of a technique and results. A technique for the local treatment of rectal cancer was performed in 29 patients who satisfied the criteria for local treatment. Four patients were classified as Dukes' stage A and 25 were stage B. Local treatment was performed by either direct excision or anorectal mobilization. The former was carried out in 13 patients in whom the tumor was located less than 7 cm from the dentate line. Excision after anorectal mobilization was performed in 16 patients with tumors located more than 7 cm from the dentate line. Patients were followed up for 5 to 10 years. They had normal rectal function after operation with normalization of rectal pressure and myoelectric activity of sphincteric muscles. Twenty four patients are alive and tumor-free. The five failures were Dukes' B category; four (30%) treated by direct excision and one (6%) by anorectal mobilization. They were subjected to a second, combined excision operation of the rectum: two patients died of distant metastasis and three are alive and tumor-free. The study showed that 12% of Dukes' B category patients had preoperative nodal deposits which escaped detection. Direct excision, owing to its high incidence of recurrence, should be restricted to Dukes' A category tumors, when the upper tumor edge is less than 7 cm from the dentate line. Otherwise, and with all Dukes' B category tumors, the anorectal mobilization operation is preferable.


Intra-luminal tumour cells and peri-anastomotic tumour growth in experimental colonic surgery. Most loco-regional tumour recurrences following colorectal cancer surgery are extra-luminal. This study was designed to determine the conditions necessary for viable intra-luminal tumour cells to give rise to extra-luminal tumour growth in an experimental animal model. When carcinoma cells were injected into the colonic lumen in the absence of a colonic wound, tumour growth was rare (1 of 24 animals). However, when tumour cells were injected intra-luminally either in the presence of an apparently watertight colotomy repair or prior to the transmural insertion of interrupted sutures, peri-anastomotic +/- widespread peritoneal tumour growth was consistently observed. Tumour growth did not occur on the mucosa of the bowel. These findings suggest that viable intra-luminal tumour cells can migrate across a clinically intact experimental large bowel anastomosis or along transmural suture tracts to give rise to extra-luminal tumour growth.


Crypt cell carcinoma of the appendix.

Crypt cell carcinoma of the appendix. A case of crypt cell carcinoma of the appendix is reported detailing its characteristic histological and immunohistochemical features and outlining current view on management.


Leiomyosarcoma of the chest wall following treatment of Hodgkin's disease. This case report describes the development of a leiomyosarcoma in the chest wall of a patient previously treated for Hodgkin's disease. Although a similar case has not been reported previously, we managed our patient with wide local excision.


Dermal angiosarcoma of the breast: a complication of primary radiotherapy? A 70-year-old female developed recurrent disease following radiotherapy for primary inoperable breast cancer 5.5 years previously. Salvage mastectomy was performed. Pathology revealed recurrent breast cancer, along with a second primary malignancy, a dermal angiosarcoma. Radical excision of recurrent angiosarcoma failed. Irradiation combined with hyperthermia showed good palliation. The angiosarcoma's possible relation to the initial radiotherapy is discussed.


Primary cystadenomatous tumors of the bile ducts. Two cases of cystadenomatous tumors of the biliary tract are presented. One was a cystadenoma and the other one was a cystadenocarcinoma. They are rare tumors, difficult to diagnose accurately preoperatively. The differential diagnosis of the cystic tumors of the liver in countries with a high incidence of liver hydatid disease is very important.


Tests of human sperm function and fertilization in vitro. To review recent studies on the development of new tests of human sperm function and evaluation of which sperm characteristics are most important for fertilization in vitro by logistic regression analysis. Recent studies on the relationship between putative and new tests of human sperm function and fertility in vitro or in vivo are discussed in this review. Some physiological and technical aspects are included. Fertilization rates in vitro and sperm tests including standard semen analysis, improved morphology assessment, objective assessment of sperm motility and movement characteristics, nuclear maturity, hypo-osmotic swelling, the acrosome and the acrosome reaction, acrosin activity, human sperm-hamster oocyte penetration assay, and sperm-zona pellucida (ZP) and sperm-oolemma binding. The percentages of sperm with normal morphology and a normal intact acrosome, mean linearity, and the number of sperm binding to the ZP were highly significant related to fertilization rates in vitro. Other sperm tests evaluated usually provided no additional information about fertilization rates. The human ZP is highly selective for binding of morphologically normal sperm. Acrosome-reacted human sperm have little or no ability to bind to the ZP. Results of in vitro fertilization can be used to evaluate tests of human sperm function. Logistic regression analysis is a powerful method for determining which groups of sperm characteristics are independently related to fertilization rates. Normal morphology, linearity, acrosome status, and sperm-ZP binding are the most important characteristics for fertilization in vitro.


Body weight, hyperinsulinemia, and gonadotropin levels in the polycystic ovarian syndrome: evidence of two distinct populations. To investigate the impact of body weight (BW) and insulin levels on gonadotropin and androgen levels in women with the polycystic ovarian syndrome (PCOS). Comparative study of endocrinologic parameters in PCOS women. University Hospital Reproductive Endocrinology Unit. Thirty obese and 19 nonobese women with PCOS. Seven obese and 7 nonobese normal women. Serum concentrations of insulin, testosterone, androstenedione, luteinizing hormone (LH), follicle-stimulating hormone. Serum LH response to gonadotropin-releasing hormone (GnRH) administration and assessment of insulin resistance by the continuous infusion of glucose with model assessment (CIGMA) test. Fasting insulin levels correlated with body mass index (BMI). Basal LH levels correlated inversely with BMI. Nonobese women with PCOS had a higher LH response to GnRH than obese women with PCOS. Only obese women with PCOS showed insulin resistance and fasting hyperinsulinemia. The data suggest that women with PCOS may be divided into two subgroups: those with obesity, insulin resistance, hyperinsulinemia, and normal/minimally elevated LH levels and those with normal BW, elevated LH levels, and normoinsulinemia.


Macroprolactinomas with suprasellar extension: effect of bromocriptine withdrawal during one or more pregnancies. To investigate the effects of bromocriptine withdrawal during one or more pregnancies in patients who presented with pituitary macroprolactinomas with suprasellar extension. Four infertile patients presenting with a macroprolactinoma with suprasellar extension conceived during treatment with bromocriptine on 10 occasions resulting in eight full-term normal deliveries. Treatment was withheld shortly after conception in each pregnancy. Serum prolactin (PRL) levels fell initially from a mean of 2,776 (range 1,682 to 4,515) to 27 micrograms/L (range 1 to 71) with the development of a partially empty sella in all patients. Recovery of visual field defects occurred in the only affected individual. In case 1, PRL levels remained within the normal range, after bromocriptine withdrawal in the first pregnancy, with the development of an empty sella. Prolactin levels, however, increased substantially in cases 2 to 4. An asymptomatic suprasellar tumor extension returned in cases 2 and 3. After two or more pregnancies (cases 1, 3, and 4), there was a progressive decline in the serum PRL levels. Although still elevated in cases 3 and 4, the PRL levels were considerably below those obtained at presentation or in the first pregnancy. Tumor regression with the development of an empty sella was observed in both these patients as well in their pregnancy or postpartum period. Bromocriptine may be safely withdrawn during pregnancy in patients presenting with a macroprolactinoma. With multiple bromocriptine induced pregnancies, PRL levels and tumor size may progressively decrease with the eventual development of an empty sella.


Relative contributions of anovulation and luteal phase defect to the reduced pregnancy rate of breastfeeding women. To evaluate the contribution of anovulation and luteal phase defects to lactational infertility. Prospective longitudinal follow-up. Outpatient clinic. Forty-nine women fully nursing and amenorrheic on day 75 postpartum and 25 cycling, interval non-nursing women. Plasma prolactin, luteinizing hormone, estradiol (E2), and progesterone (P) levels twice a week up to the second postpartum menses. Ovulation rate and endocrine profile of the menstrual cycles. Ovulation rates were 37% and 97% at 6 and 12 months postpartum; 67% of ovulations occurred in amenorrhea. The luteal phase was shorter, and E2 and P levels were lower in lactating women than in non-nursing women. These parameters were closer to normal in the second cycle than the first, in spite of active nursing. The risk of ovulation and pregnancy in amenorrhea was 27.7% and 0.9% at month 6 postpartum. After the first menses, these risks were 93% and 7%, respectively. The abnormal endocrine profile of the first luteal phase offers effective protection to women who ovulate during lactational amenorrhea within the first 6 months after delivery. Later luteal phases are improved and women are at risk of pregnancy.


Effects of clomiphene citrate on pituitary luteinizing hormone and follicle-stimulating hormone release in women before and after treatment with ethinyl estradiol. To evaluate the effects of clomiphene citrate (CC) on pituitary luteinizing hormone (LH) and follicle-stimulating hormone (FSH) release in hypoestrogenic women and in the same women after treatment with ethinyl estradiol (EE2). The study was of a prospective nature and was conducted on selected patients. Volunteer women were studied in a tertiary care public hospital. The 10 patients studied were selected on the basis of hypogonadal status (menopause, premature ovarian failure, or gonadal dysgenesis and Turner phenotype) and no hormonal treatment. Gonadotropin-releasing hormone (GnRH) was continually infused at the dose of 0.2 micrograms/min for 4 hours before and after the use of CC and/or EE2. The study was performed with the objective of determining the effect of estrogen (E) levels on the action of CC on in vivo gonadotropin release. In the presence of hypoestrogenic conditions, CC had no pituitary action. However, after EE2 treatment CC promoted greater FSH release and a significant inhibition of LH release from the pituitary. Clomiphene citrate needs a basal E level to be able to act on the pituitary. In normoestrogenic states and under GnRH stimulation, CC preferentially promotes FSH release while presenting a predominantly inhibitory effect on LH release.


Hysteroscopic findings after missed abortion. To identify the relation of missed abortions and intrauterine pathology. A postabortal hysteroscopy was performed 8 to 12 weeks after a dilatation and curettage (D&C) for missed abortion. Department of Obstetrics and Gynecology, Assaf Harofeh Medical Center, Zerifin, Israel. Sixty patients after a D&C for a missed abortion. The hysteroscopic appearance of the uterine cavity. Intrauterine adhesions occurred in only 10 patients (16.7%) with most of them of the mild type. A previous missed abortion was recorded in 60% of these cases. Uterine anomaly was found in one quarter of the cases, mainly an incomplete uterine septum. We believe that missed abortion does not predispose for intrauterine adhesions to the extent that was previously believed. A partial uterine septum is a major factor predisposing for the occurrence of missed abortion. Hysteroscopy after a missed abortion is an easy and efficient means for both identifying such uterine anomaly and excluding intrauterine adhesions.


Detection of antiendometrial antibodies in patients with endometriosis: methodological issues. To determine if methodological reasons explain previous inability of a single investigator to detect antiendometrial antibodies in patients with endometriosis by indirect immunofluorescence (IIF). Sera of selected patients were tested by standard IIF and recently reported methodology. Alternate diluent experiments were conducted to explain discordant results. Sera were obtained preoperatively, and testing was completed in an institutional research laboratory. Results are compared with surgical findings. Patients, seven with endometriosis and six without endometriosis, 40 years of age or less who are presenting for gynecological surgery. None. Indirect immunofluorescence was graded according to the presence or absence of cytoplasmic staining, intensity of the staining, and the lowest dilution at which staining was observed. The standard IIF method repeatedly demonstrated the presence of cytoplasmic staining for the presence of antiendometrial antibodies in patients with endometriosis, whereas modified methods did not produce similar positive staining results. During the alternate dilution experiments, when fluorescein isothiocyanate conjugate (FITC) was diluted in animal sera, negative results were obtained. The variations used in alternate methodology and failure to titrate FITC reported by other investigators are possible reasons for false-negative results reported.


An office semiquantitative serum human chorionic gonadotropin determination. To compare the human chorionic gonadotropin (hCG) concentration, established by a standard serum quantitative hCG fluorescent immunoassay and a semiquantitative serum determination. Patients were asked to give two serum samples early in pregnancy to establish the accuracy of a semiquantitative serum hCG test in diluted and undiluted serum samples. In a laboratory setting, two serum samples were determined; one sample was submitted for standard serum hCG radioimmunoassay, and the other was tested for hCG by the 25 IU Tandem ICON Assay (Hybritech, Liege, Belgium) in diluted and undiluted serum samples. Sixteen patients supposed to be pregnant. Within dilutional zones, the results of a semiquantitative hCG test were compared with a known standard quantitative serum hCG immunoassay measurements. The semiquantitative hCG ranges of serum hCG compare fairly well with an accurate standard quantitative serum hCG immunoassay. The determination of a serum hCG range compares well with the standard quantitative serum hCG immunoassay (First International Reference Preparation) and can be completed within 15 minutes. This office semiquantitative serum hCG determination proved to be a quick and reliable test.


Effects of melatonin on progesterone production by human granulosa lutein cells in culture. To test the hypothesis that melatonin modulates steroid synthesis in the human ovary. Granulosa lutein cells obtained from in vitro fertilization cycles were cultured in medium containing melatonin and human chorionic gonadotropin (hCG). Progesterone (P) secretion by granulosa lutein cells increased progressively in both basal and hCG-stimulated conditions, up to 96 hours in culture, plateaued at 144 and decreased thereafter. Melatonin (10(-7), 10(-9), 10(-11) M) had no effect on basal P or 17 beta-estradiol production. The addition of melatonin to the hCG-treated granulosa lutein cells significantly (P less than 0.05) potentiated the stimulatory effect of hCG on P production. The effect was most prominent after 144 and 196 hours of incubation. This observation suggests a role for melatonin in the intraovarian control of P production in the human ovary.


Estradiol stimulates the secretion of prostacyclin and thromboxane from endometrial stromal cells in culture. To determine the production of the vasoactive prostaglandins (PGs), prostacyclin, and thromboxane (TX) in endometrial stromal cell cultures and their steroid modulation by sex steroids. Primary stromal cell cultures with steroid modulation in tissues from the follicular and luteal phases of normal women. Laboratories of the Division of Reproductive Endocrinology and tissues obtained from normal patients at the Los Angeles County and University of Southern California Medical Center in Los Angeles, California. Stromal cell tissue culture for 14 days. Radioimmunoassay of 6-keto-PGF1 alpha, TXB2, and prolactin (PRL) as well as determination of protein content. Estradiol stimulated 6-keto-PGF1 alpha production more than TXB2. This effect occurred after day 6 and was as much as 10-fold greater in luteal phase tissue. The effect was eliminated by progestins that alone had no effect. This modulatory response was greater in luteal endometrial tissue than in follicular phase tissue. As expected, in these tissues progestin stimulated PRL, an effect opposite to its negative influence on the vasoactive PGs. These data provide evidence for a steroid-related influence on the balance of vasoactive PGs in the stroma of the endometrium.