Constitutive secretion of a bacterial enzyme by polarized epithelial cells. The constitutive (or default) pathway for protein secretion was investigated in two epithelial cells, Madin-Darby canine kidney (MDCK) and human colonic adenocarcinoma (Caco-2), using a bacterial enzyme. The choice of a bacterial protein was based on the requirement to identify a protein devoid of sorting signals. The sorting of a bacterial endoglucanase derived from Clostridium thermocellum, endoglucanase E, from stably transfected MDCK and Caco-2 cells was examined. The choice of a bacterial endoglucanase for these studies has advantages of simple, sensitive and quantitative detection, while higher eukaryotic cells do not express endoglucanase activity. Both cell lines secreted a 50 kDa form of the bacterial protein, while smaller intracellular forms were also observed. In polarized layers of MDCK cells the endoglucanase was secreted into both membrane domains in the ratio 62% apical and 38% basolateral. In Caco-2 cells secretion was predominantly, 70%, through the basolateral membrane. These results define the constitutive pathway for protein secretion in these two model epithelial cells.


Effect of caffeine on intracellular transport of Semliki Forest virus membrane glycoproteins. The effect of caffeine on the intracellular transport of Semliki Forest virus (SFV) membrane glycoproteins was studied in baby hamster kidney (BHK) cells. The movement of the proteins was affected at two steps in the exocytic pathway. The exit of the proteins from the endoplasmic reticulum (ER) was inhibited by 10 mM caffeine at 20 degrees C, a temperature that normally allows transport to the Golgi complex. At higher temperatures (28 degrees C and 37 degrees C) in the presence of 10 mM caffeine exit from the ER occurred, but the proteins accumulated at intracellular membrane elements. Immunofluorescence localization, endoglycosidase-H analysis, and analysis of the proteolytical cleavage of the p62 precursor protein suggested that transport in the presence of 10 mM caffeine was arrested at the membranes between the trans-Golgi and the plasma membrane.


Origins of the parasitophorous vacuole membrane of the malaria parasite, Plasmodium falciparum, in human red blood cells. We have attempted to determine whether the parasitophorous vacuole membrane, in which the malaria parasite (merozoite) encapsulates itself when it enters a red blood cell, is derived from the host cell plasma membrane, as the appearance of the invasion process in the electron microscope has been taken to suggest, or from lipid material stored in the merozoite. We have incorporated into the red cell membrane a haptenic phospholipid, phosphatidylethanolamine, containing an NBD (N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)) group, substituted in the acyl chain, and allowed it to translocate into the inner bilayer leaflet. After invasion of these labelled cells by the parasite, Plasmodium falciparum, immuno-gold electron microscopy was used to follow the distribution of the labelled lipid; this was found to be overwhelmingly in favour of the host cell membrane relative to the parasitophorous vacuole. Merozoites of P. knowlesi were allowed to attach irreversibly to red cells without invasion, using the method of pretreatment with cytochalasin. The region of contact between the merozoite and the host cell membrane was in all cases devoid of the labelled phosphatidylethanolamine. These results lead us to infer that the parasitophorous vacuole membrane is derived wholly or partly from lipid preexisting in the merozoite.


Syringe loading introduces macromolecules into living mammalian cell cytosol. We describe a simple, efficient, gentle and inexpensive technique for the introduction of normally impermeant macromolecules into the cytosol of living mammalian cells growing in suspension or attached to the culturing substratum. Loading is achieved by the production of transient, survivable plasma membrane disruptions as cells are passed back and forth through a standard syringe needle or similar narrow orifice. The loading volume required, which contains cells and the macromolecule to be loaded, can be as little as 5 microliters, thus minimizing the use of valuable reagents. In addition, we report that the surfactant molecule, Pluronic F-68, is capable of altering the physical properties of the plasma membrane in such a way as to increase loading efficiency and the long-term survivability of cells loaded by this and other mechanically based cell-loading techniques.


Structure of the cortical cytoskeleton in mammalian outer hair cells. The cortical cytoskeletal lattice in outer hair cells is a two-dimensional actin-based structure, which can be labelled with rhodamine/phalloidin and disrupted by the enzyme deoxyribonuclease I. Structural information from thin sectioned, freeze-etched and negatively stained preparations shows that it is based upon two types of filament that form a cross-linked lattice of circumferential filaments. The cross-links are 70-80 nm long. Measurements of the spacing between circumferential filaments suggest that the lattice is stiffer circumferentially than it is longitudinally. Analysis of the orientation of circumferential filaments shows that it is composed of discrete domains of up to 10 microns 2. Relative movements between domains could allow substantial changes of cell shape without disrupting the unit structure of the lattice, thus allowing the cell cortex to retain its elastic responses to high-frequency deformations.


Characterization of the enterocyte-like brush border cytoskeleton of the C2BBe clones of the human intestinal cell line, Caco-2. The brush border (BB) of the enterocyte is a well-studied example of the actin-based cytoskeleton. We describe here a cell culture model that expresses a faithful representation of the in vivo structure. Two clones (C2BBe 1 and 2) isolated from the cell line Caco-2 (derived from a human colonic adenocarcinoma) formed a polarized monolayer with an apical BB morphologically comparable to that of the human colon. BBs could be isolated by standard methods and contained the microvillar proteins villin, fimbrin, sucrase-isomaltase and BB myosin I, and the terminal web proteins fodrin and myosin II. The immunolocalization of these proteins in confluent, filter-grown monolayers was determined by laser scanning confocal microscopy; patterns of distribution comparable to those in human enterocytes were observed. Sedimentation analysis of cell homogenates derived from C2BBe cells and human colonic epithelial cells demonstrated similar patterns of fractionation of BB proteins; the physical association of those proteins, as determined by extraction from the BB, was also comparable between the two cell types. Like enterocytes of the human intestine, C2BBe cells expressed multiple myosin I immunogens reactive with a head domain-specific monoclonal antibody raised against avian BB myosin I, one of which co-migrated with the approximately 110 kilodalton (kDa) heavy chain of human BB myosin I. In addition, the C2BBe cells express a pair of higher molecular mass immunogens (130 and 140 kDa). These myosin I immunogens all exhibit ATP-dependent association with the C2BBe cytoskeleton. Although the higher molecular mass immunogens were detected in several other human intestinal lines examined, including the parent Caco-2 line, none of these other lines expressed detectable levels of the 110 kDa immunogen, which is presumed to be the heavy chain of human BB myosin I.


Stage-specific tyrosine phosphorylation of actin in Dictyostelium discoideum cells. A 45 kDa protein in Dictyostelium discoideum cells that was recognized by a phosphotyrosine-specific antibody was identified by its binding activity to DNase I and its 2D-electrophoretic behavior as actin. The reactivity of actin with the antibody was transiently enhanced for about 30 minutes shortly after starving cells were reintroduced into nutrient medium. This effect indicates a modification of actin that is regulated under physiological conditions. A similar effect was obtained when growing cells were treated with phenylarsine oxide (PAO), an inhibitor of phosphotyrosine phosphatases. This effect was reversed and the cells fully recovered upon addition of the PAO antagonist 2,3-dimercaptopropanol. Starved cells did not show this enhancement of antibody labelling, which indicates that the response to PAO depends on the developmental stage. Phosphorylated amino acid residues were identified after in vivo labelling with [32P]phosphate in the presence of PAO. Part of the radioactivity in the actin band was recovered as phosphotyrosine, another part as phosphoserine. PAO caused the cells to form elongated blebs, to round up and finally to become immobilized. Fluorescence labelling with phalloidin of cells that were fixed at different times of PAO treatment revealed a progressive decrease in the staining for actin filaments and showed that these alterations in cytoskeleton organization were readily reversible, in accordance with the reversal of tyrosine phosphorylation at actin.


The localization of myosin I and myosin II in Acanthamoeba by fluorescence microscopy. We used several fixation protocols and a panel of monoclonal antibodies to re-examine the localization of myosin I and myosin II in Acanthamoeba. Two monoclonal antibodies that bind to the head of myosin II stain a range of particles in the cytoplasm. The smallest and most numerous cytoplasmic particles are about the same size and intensity as myosin II minifilaments and are distributed throughout the endoplasm. The largest particles stain like myosin II thick filaments and are concentrated in the cleavage furrow of dividing cells and in the tail of locomoting cells. Five different monoclonal antibodies that bind to the myosin II tail also stain cytoplasmic particles but with a limited range of intensity. None of the myosin II monoclonal antibodies stains the contractile vacuole or plasma membrane. Two monoclonal antibodies to myosin I gave punctate cytoplasmic staining that did not correspond clearly to any of the phase-dense particles in the cytoplasm. In many, but not all, locomoting cells, the myosin I staining was concentrated at the leading edge. Both myosin I antibodies stained a single cytoplasmic vacuole of variable size that was presumed to be the contractile vacuole. The antibody that binds myosin IA but not myosin IB stained novel intercellular contacts and the antibody that binds both myosin IA and myosin IB stained the plasma membrane, especially the tips of filopodia.


Formation of highly organized skeletal muscle fibers in vitro. Comparison with muscle development in vivo. Two methods were developed in which long-term cultures of quail skeletal muscle were established so that all of the muscle fibers develop in a highly oriented manner. The muscle fibers became spontaneously and vigorously contractile and established strong connections with the extracellular matrix at their ends that closely duplicate the structure of the myotendinous junction. A continuous basal lamina was formed around each muscle fiber that contained type IV collagen, laminin and heparan sulfate proteoglycan. With one of the methods, an extensive extracellular matrix developed around each muscle fiber that was highly organized with the formation of a distinctive epimysium, perimysium and endomysium. Analysis of the cultures by both methods for different isoforms of myosin showed expression of an adult form of myosin by some of the muscle cells. The results therefore demonstrate that muscle development in the present culture systems proceeds extensively for several weeks. It will now be possible to investigate directly the structure of the connections between muscle fibers and the extracellular matrix.


Neuroreceptor quantitation in vivo by the steady-state principle using constant infusion or bolus injection of radioactive tracers. The approaches hitherto used for measuring the kinetic constants Kd and Bmax of neuroreceptors in vivo all violate the steady state of the system. This complicates the kinetic analysis as approximations must be made, introducing errors of unknown magnitude. The present study presents the theory for designing experiments in which the steady state is preserved. It is based on maintaining a constant degree of receptor binding (occupancy) throughout the experiment. This is achieved by administering by prolonged intravenous infusion the non-radioactive ligand one wishes to study. The fraction of receptors sites not occupied by the "cold" ligand is measured by using trace amounts of a radioactive ligand binding to the same receptor. A minimum of two studies at different occupanies must be performed. In this presentation it is proposed to make the second study at essentially zero receptor occupancy by administering the tracer alone. The pair of tracer studies, the one without and the other with infusion of cold ligand, allows calculation of the cold ligand's equilibrium dissociation constant Kd. In the special case when tracer and cold ligands are chemically identical, then Bmax can also be calculated. Two different modes of tracer administration can be used. If the tracer is also infused at a constant rate for a long time, then the occupancy of receptor sites by the cold ligand can be calculated by measuring the equilibrium tracer concentrations in brain and plasma. If the tracer is administered as an intravenous bolus injection, then the area under the brain and plasma radioactivity curves or compartmental analysis must be used.(ABSTRACT TRUNCATED AT 250 WORDS)


Repeated negative DC deflections in rat cortex following middle cerebral artery occlusion are abolished by MK-801: effect on volume of ischemic injury. Following permanent occlusion of the left middle cerebral artery (MCA) in rats, electrophysiological and hemodynamic characteristics of the periinfarct border zone were investigated in sham-operated (n = 6), untreated (n = 6), and MK-801-treated (3.0 mg/kg; n = 6) animals. For this purpose, direct current potential (DC), EEG, and blood flow (laser-Doppler flowmetry) were recorded from the cortex in the periphery of the MCA territory. In sham-operated rats, a single negative cortical DC deflection was observed after electrocoagulation of the cortex, whereas in untreated MCA-occluded animals, three to eight transient DC deflections were monitored during the initial 3 h of ischemia. The duration of these cortical DC shifts gradually increased from 1.2 +/- 0.3 to 3.7 +/- 2.7 min (mean +/- SD; p less than 0.05) during this time. In animals treated intraperitoneally with MK-801 (3.0 mg/kg) immediately after MCA occlusion, the number of cortical DC shifts significantly declined to one to three deflections (p less than 0.005). The EEG of the treated animals revealed low-amplitude burst-suppression activity. In the untreated and treated experimental group, the reduction of cortical blood flow amounted to 69 +/- 25 and 49 +/- 13% of control, respectively. Despite the more pronounced cortical oligemia, MK-801 treatment resulted in a significant decrease of the volume of the ischemically injured tissue from 108 +/- 38.5 (untreated group) to 58 +/- 11.5 (p less than 0.05) mm3. Our results suggest that repetitive cortical DC deflections in the periinfarct border zone contribute to the expansion of ischemic brain infarcts.


Elevated lactate and alkalosis in chronic human brain infarction observed by 1H and 31P MR spectroscopic imaging. The goal of this study was to investigate lactate and pH distributions in subacutely and chronically infarcted human brains. Magnetic resonance spectroscopic imaging (MRSI) was used to map spatial distributions of 1H and 31P metabolites in 11 nonhemorrhagic subacute to chronic cerebral infarction patients and 11 controls. All six infarcts containing lactate were alkalotic (pHi = 7.20 +/- 0.04 vs. 7.05 +/- 0.01 contralateral, p less than 0.01). This finding of elevated lactate and alkalosis in chronic infarctions does not support the presence of chronic ischemia; however, it is consistent with the presence of phagocytic cells, gliosis, altered buffering mechanisms, and/or luxury perfusion. Total 1H and 31P metabolites were markedly reduced (about 50% on average) in subacute and chronic brain infarctions (p less than 0.01), and N-acetyl aspartate (NAA) was reduced more (approximately 75%) than other metabolites (p less than 0.01). Because NAA is localized in neurons, selective NAA reduction is consistent with pathological findings of a greater loss of neurons than glial cells in chronic infarctions.


Increased cerebral blood flow in anemic patients on long-term hemodialytic treatment. CBF was measured in 15 patients on chronic hemodialytic treatment. CBF was measured with xenon-133 inhalation using single photon emission tomography. In addition, computerized tomography (CT) and a neurological examination were done prior to hemodialysis. Mean CBF was 66.2 +/- 17.3 (SD) ml 100 g-1 min-1, which was significantly higher (t-test, p less than 0.05) than for an age-matched control group (54.7 +/- 10.2 ml 100 g-1 min-1). However, the hematocrit for the patients was considerably lower, 0.30 +/- 0.07, as compared to 0.43 +/- 0.03 in the controls. A significant negative correlation was observed between CBF and the hematocrit (y = -1.79x + 120.7, r = -0.71, p less than 0.01). Calculating CBF from this equation in the dialyzed patients using a hematocrit of 0.43 yielded a mean CBF value of 43.7 ml 100 g-1 min-1, i.e., 20% below the expected. Two patients showed a focal CBF decrease. CT showed central or cortical atrophy in five patients, and two had small hypodense lesions. The neurological examination revealed slight to moderate dementia in seven cases. Although mean CBF was found to be increased by 21% as compared to the control group, an even higher CBF level would have been expected to outweigh the decreased oxygen carrying capacity of the blood. The findings suggest a lowered metabolic demand of the brain tissue, probably due to subtle brain damage.


Distributions of heat shock protein-70 mRNAs and heat shock cognate protein-70 mRNAs after transient global ischemia in gerbil brain. Distributions of heat shock protein (HSP)-70 mRNAs and heat shock cognate protein (HSC)-70 mRNAs after 10 min of transient global ischemia were investigated in gerbil forebrain by in situ hybridization using cloned cDNA probes selective for the mRNAs. Expression of HSP70 immunoreactivity was also examined in the same brains. In hippocampal CA1 neuronal cells, in which only a minimal induction of immunoreactive HSP70 protein was found, the strong hybridization for HSP70 mRNA disappeared at around 2 days before the death of CA1 cells became evident. Furthermore, in hippocampal CA3 cells, a striking induction of HSP70 mRNA was sustained even at 2 days along with a prominent accumulation of HSP70 immunoreactivity. In contrast to the case of HSP70 mRNA, HSC70 mRNA was present in most neuronal cells, especially dense in CA3 cells, of the sham brain. A co-induction of HSP70 and HSC70 mRNAs was observed in several cell populations after the reperfusion with a peak at 8 h, although the magnitude of HSC70 mRNA induction was lower than that of HSP70 mRNA, particularly in CA1 cells. The expression of HSC70 mRNA in CA1 cells also disappeared at around 2 days. All the induced signals of HSP70 and HSC70 mRNAs in other cell populations were diminished and returned to the sham level, respectively, by 7 days. These results are the first to show the time courses of distribution of HSP70 and HSC70 mRNAs and the immunoreactive HSP70 protein in the same gerbil brain after ischemia.(ABSTRACT TRUNCATED AT 250 WORDS)


Uncoupled cerebral blood flow and metabolism after severe global ischemia in rats. In a rat model of complete global brain ischemia (neck tourniquet) lasting either 3 min or 20 min, we monitored global CBF (sagittal sinus H2 clearance) and CMRO2 for 6 h to test the hypothesis that delayed postischemic hyperemia and uncoupling of CBF and CMRO2 occur depending on the severity of the insult. Early postischemic hyperemia occurred in both the 3-min and 20-min groups (p less than 0.05 vs. baseline values) and resolved by 15 min. Hypoperfusion occurred in the 3-min group between 15 and 60 min postischemia (approximately 23% reduction), and in the 20-min group from 15 to 120 min postischemia (approximately 50% reduction) (p less than 0.05), and then resolved. CMRO2 was not significantly different from baseline at any time after ischemia in the 3-min group. After 20 min of ischemia, however, CMRO2 was decreased (approximately 60%) throughout the postischemic period (p less than 0.05). At 5 min after ischemia, CBF/CMRO2 was increased in both groups but returned to baseline from 60 to 120 min postischemia. In the 3-min group, CBF/CMRO2 remained at baseline throughout the rest of the experiment. However, in the 20-min group, CBF/CMRO2 once again increased (approximately 100%), reaching a significant level at 180 min and remaining so for the rest of the 6-h period (p less than 0.05). These data demonstrate biphasic uncoupling of CBF and CMRO2 after severe (20 min) global ischemia in rats. This relatively early reemergence of CBF/CMRO2 uncoupling after 180 min of reperfusion is similar to that observed after prolonged cardiac arrest and resuscitation in humans.


Methodological requirements for accurate measurements of brain and body temperature during global forebrain ischemia of rat. The methodological requirements for accurate measurements of brain and body temperature during brain ischemia have been validated in Wistar rats submitted to 30 min of four-vessel occlusion. During ischemia, brains were exposed to three different temperature profiles: spontaneous cooling from 36 to 31 degrees C (n = 10), constant hypothermia at 30 degrees C (n = 19), and constant normothermia at 36 degrees C (n = 21). Direct and indirect brain temperature recordings were carried out by placing fine thermocouples (200 microns diameter) into the striate nucleus, the temporal muscle, and the epidural space. Body temperature was measured with a flexible thermocouple inserted at various depths into the rectum. Accurate measurements of body temperature required insertion of the rectal probe to a depth of at least 6 cm; lesser insertion resulted in an underestimation of up to 6 degrees C. Accurate estimates of brain temperature were obtained in all three experimental conditions by recording of the epidural temperature. The temperature in the temporal muscle, by contrast, differed from the brain temperature by up to 2 degrees C, depending upon the experimental condition and the duration of ischemia. We therefore suggest that indirect measurements of brain temperature during ischemia are carried out in the epidural space in order to avoid misinterpretations of temperature-sensitive pathological changes.


Influence of preischemic hyperglycemia on osmolality and early postischemic edema in the rat brain. Preischemic hyperglycemia, which raises tissue lactate content during ischemia, is known to aggravate ischemic brain damage. To explore the possibility that the enhanced lactic acidosis gives rise to osmotic damage, we studied the influence of a varied preischemic plasma glucose concentration on the early postischemic edema. Brain edema was measured by the specific-gravity technique. Brain and plasma osmolality were measured with a vapor pressure osmometer. We examined different brain regions in hyperglycemic and moderately hypoglycemic rats subjected to 15 min of forebrain ischemia, followed by recirculation for 5, 15, and 30 min. The decrease in specific gravity was compared with the increase in osmolality, to study whether the edema formation in the different groups correlated to the increase in tissue osmolality. We found edema formation to be most pronounced in frontoparietal cortex. In this structure and in hippocampus, statistically significant decreases of specific gravity were seen at all recirculation times studied. In caudoputamen, significant edema was seen only in the groups with 5 and 15 min of recirculation. Contrary to expectations, no difference was found between hyperglycemic and hyperglycemic animals. Tissue osmolality increased during ischemia in both the low and high glucose groups, but to a higher level in the latter (hypoglycemia 311 +/- 1 mmol kg-1, hyperglycemia 328 +/- 10 mmol kg-1; mean +/- SD, p less than 0.05). In the hyperglycemic group, brain osmolality remained elevated for the first 15 min of recirculation.(ABSTRACT TRUNCATED AT 250 WORDS)


Errors introduced by tissue heterogeneity in estimation of local cerebral glucose utilization with current kinetic models of the [18F]fluorodeoxyglucose method. The effects of tissue heterogeneity on the estimation of regional cerebral glucose utilization (rCMRglc) in normal humans with [18F]2-fluoro-2-deoxy-D-glucose ([18F]FDG) and positron emission tomography (PET) were compared with respect to the various kinetic models of the [18F]FDG method. The kinetic models were conventional homogeneous tissue models of the [18F]FDG method, with (4K Model) and without (3K Model) a rate constant to account for an apparent loss of [18F]2-fluoro-2-deoxy-D-glucose-6-phosphate ([18F]FDG-6-P), and a tissue heterogeneity model (TH Model). When either of the kinetic models designed for homogeneous tissues was applied to heterogeneous tissues, estimates of the rate constant for efflux of [18F]FDG from the tissue (k2*) and of the rate constant for phosphorylation of [18F]FDG (k3*) decreased as the duration of the experimental period was increased. When the 4K Model was used, estimates of the rate constant for the apparent dephosphorylation of [18F]FDG-6-P (k4*) were significantly greater than zero and fell with increasing duration of the experimental period. Although the TH Model included no term to describe an apparent dephosphorylation of [18F]FDG-6-P, the fit of the TH Model to the time course of total tissue radioactivity was at least as good as and often better than the fit of the 4K Model in the 120-min period following the pulse of [18F]FDG. Hence, the high estimates of k4* found in PET studies of less than or equal to 120 min can be explained as the consequence of measuring radioactivity in a heterogeneous tissue and applying a model designed for a homogeneous tissue; there remains no evidence of significant dephosphorylation of [18F]FDG-6-P in this time period. Furthermore, use of the 4K Model led to an overestimation of rCMRglc; whole-brain glucose utilization calculated with the 4K Model was greater than 20% higher than values usually obtained in normal humans by the model-independent Kety-Schmidt technique. rCMRglc was accurately estimated by the TH Model and, in experimental periods sufficiently long to minimize the effects of tissue heterogeneity, also by the original 3K Model of the deoxyglucose method.


Sciatic nerve stimulation does not increase endogenous adenosine production in sensory-motor cortex. Adenosine participates in the coupling of cerebral blood flow to oxygen consumption in the brain during such stimuli as hypoxia, ischemia, and seizures. It has been suggested that it also participates in the regulation of cerebral blood flow during somatosensory stimulation, a condition during which cerebral blood flow and oxygen consumption appear to be uncoupled. Interstitial adenosine was estimated by the microdialysis technique and cerebral blood flow was measured by hydrogen clearance in the hindlimb sensory-motor cortex during sciatic nerve stimulation. Cerebral blood flow increased from 102 to 188 ml min-1 100 g-1 (p less than 0.001) in the cortex contralateral to the stimulated leg without an associated increase in interstitial adenosine (baseline 0.624 microM, stimulation 0.583 microM). Infusion of the adenosine antagonist 8-sulfophenyltheophylline failed to block an increase in cerebral blood flow during central sciatic nerve stimulation, but decreased basal cerebral blood flow (69 ml min-1 100 g-1). These results suggest that adenosine does not mediate changes in cerebral blood flow during somatosensory stimulation, but may participate in the regulation of cerebral blood flow in the basal state.


Stimulation of Cl area neurons globally increases regional cerebral blood flow but not metabolism. We examined the effects of electrical and chemical stimulation of the Cl area of the rostral ventrolateral medulla (RVL) on regional cerebral blood flow (rCBF) and regional cerebral glucose utilization (rCGU) in anesthetized (chloralose), paralyzed (curare) and ventilated rats. rCBF and rCGU were measured using 14C-iodoantipyrine (IAP) and 14C-deoxyglucose (2-DG), respectively, as indicators, with bilateral regional dissection of 11 brain regions. Electrical stimulation of the RVL elicited increases in arterial pressure (AP), heart rate (HR) and plasma concentration of epinephrine (EPI) and norepinephrine (NE). In addition, stimulation of the RVL, but not the adjacent medial longitudinal fasciculus, with AP maintained, increased rCBF (p less than 0.05, n = 6), but not rCGU, bilaterally and symmetrically (134-169% of control) throughout the brain. Bilateral adrenalectomy abolished the increase in plasma EPI elicited by stimulation of the RVL but did not affect resting rCBF (n = 5) or the elevation in rCBF elicited by RVL stimulation (n = 5). Increases in rCBF elicited by RVL stimulation were also unaffected by acute transection of the superior cervical ganglion (p greater than 0.05). Kainic acid (KA) microinjected into the RVL unilaterally (n = 6) at a dose producing sustained elevation in AP (5 nmol in 100 nl), elicited changes in rCBF similar to those elicited by electrical stimulation. We conclude that neurons within the RVL, possibly those of the adrenergic Cl group, can initiate a global cerebrovasodilation, but not an increase in rCGU, largely through neural pathways intrinsic to the brain. The responses may represent activation of networks in RVL mediating circulatory adjustments to hypoxia.


Regulation of cerebral glucose metabolism in normal and polycythemic newborn lambs. In contrast to previous investigations, a recent study of polycythemic lambs suggested that cerebral glucose delivery (concentration x blood flow), not arterial glucose concentration, determined cerebral glucose uptake. In the present study, the independent effects of arterial glucose concentration and delivery on cerebral glucose uptake were examined in two groups of chronically catheterized newborn lambs (control and polycythemic). Arterial glucose concentration was varied by an infusion of insulin. CBF was reduced in one group of lambs (polycythemic) by increasing the hematocrit. At all arterial glucose concentrations, the cerebral glucose delivery of the polycythemic group was 59.6% of the control group. At arterial glucose concentrations of greater than 1.6 mmol/L, cerebral glucose uptake was constant and similar in both groups. At arterial glucose concentrations of less than or equal to 1.6 mmol/L, cerebral glucose uptake was unchanged in the control group, but was significantly decreased in the polycythemic group. In contrast, the cerebral glucose uptake was similar in both groups over a broad range of cerebral glucose delivery values. At cerebral glucose delivery values less than or equal to 83 mumols/min/100 g, there was a significant decrease in cerebral glucose uptake in both groups. During periods of low cerebral glucose delivery and uptake, cerebral oxygen uptake fell in the control group but remained unchanged in the polycythemic group. Maintenance of cerebral oxygen uptake in the polycythemic group was associated with an increased extraction and uptake of lactate and beta-hydroxybutyrate. We conclude that cerebral glucose delivery, not arterial glucose concentration alone, determines cerebral glucose uptake.


Effects of indomethacin on myogenic contractile activation and responses to changes in O2 and CO2 in isolated feline cerebral arteries. We used an isolated, pressurized, and perfused feline middle cerebral artery preparation to measure how changes in intraluminal pressure and alterations in O2 and CO2 affect vessel diameter and myogenic contractile activation before and after treatment with indomethacin (IND). Vessel diameters were measured over the pressure range 60-140 mm Hg. The arteries were then exposed to low O2 (50 torr) and/or high CO2 (65 torr) and diameters remeasured over the same range. Under control conditions, the arteries exhibited myogenic contractile activation. Exposure to low O2, high CO2, or a mixture of low O2/high CO2, increased vessel diameter but did not change the vessels' myogenic contractile responsiveness to changes in pressure. Arteries exposed to IND decreased in diameter but retained myogenic contractile activity. In the presence of IND, vessels dilated to both low O2 and a mixture of low O2/high CO2, but did not dilate to high CO2 alone. Under all conditions, vessels retained myogenic contractile activity. Results obtained under control conditions and low O2 confirm those of others using similar systems. Myogenic contractile activity in the presence of high CO2 or a mixture of low O2/high CO2 has not been previously reported. The dilation to low O2 but not to high CO2 in the presence of IND suggests that this drug's effects in cerebral arteries are not limited solely to inhibition of prostaglandin synthesis.


Intracerebral microdialysis of extracellular amino acids in the human epileptic focus. Extracellular levels of aspartate (ASP), glutamate (GLU), serine (SER), asparagine (ASN), glycine (GLY), threonine (THR), arginine (ARG), alanine (ALA), taurine (TAU), tyrosine (TYR), phenylalanine (PHE), isoleucine (ILEU), and leucine (LEU) were monitored by using intracerebral microdialysis in seven patients with medically intractable epilepsy, undergoing epilepsy surgery. In association with focal seizures, dramatic increases of the extracellular ASP, GLU, GLY, and SER concentrations were observed. The other amino acids analyzed, including TAU, showed small changes. The results support the hypothesis that ASP, GLU, GLY, and possibly SER, play an important role in the mechanism of seizure activity and seizure-related brain damage in the human epileptic focus.


The effect of profound levels of hypothermia (below 14 degrees C) on canine cerebral metabolism. The goal of this study was to determine the temperature coefficient (Q10) for canine CMRO2 at temperatures below 14 degrees C. Eight dogs were anesthetized with halothane for surgical preparation. The animals were placed on total cardiopulmonary bypass and CBF was measured by direct sagittal sinus outflow. Duplicate measurements were taken at 37, 13, and 7 degrees C. The EEG became isoelectric at a temperature of 12.0 +/- 0.8 degrees C. The Q10 between 13 and 7 degrees C was 2.19 +/- 0.59. With rewarming to 37 degrees C, cerebral metabolic variables returned to control levels. Brain biopsies taken at the end of the study yielded normal values for brain energy stores. We conclude that the Q10 for CMRO2 at temperatures between 7 and 37 degrees C can be profoundly affected by the state of cerebral function as reflected by the EEG. In the absence of EEG activity, an expected Q10 value of 2.2 reflects only the direct effect of temperature on the rates of biologic reactions.


Plasma L-[18F]6-fluorodopa input function: a simplified method. This article describes a simplified method for the determination of the L-[18F]6-fluorodopa (FDOPA) fraction time course that takes advantage of the strong correlation between the radioactivity ratio (metabolites/FDOPA) and time. Serial arterial blood samples are collected for assay of plasma total radioactivities following an intravenous injection of FDOPA into carbidopapretreated subjects. In addition, a single plasma sample, collected late in the study and analyzed for FDOPA fraction, is sufficient to determine accurately the time course of the FDOPA concentration in plasma. The validated straight-line method greatly simplifies blood analysis for routine positron emission tomography FDOPA studies.


Enzyme-linked immunosorbent assay for the phytotoxin cleistanthin A. Enzyme-linked immunosorbent assay is reported for the estimation of cleistanthin A, a major constituent of the toxic plant Cleistanthus collinus. Rabbit antibodies were obtained by immunisation with cleistanthin A hemisuccinate-BSA conjugate and the ELISA developed thereupon could detect cleistanthin A at as low a concentration as 3 ng/ml. Cross-reactivity studies with structural analogs as well as with other phytotoxins and drugs of common occurrence established the suitability of the ELISA to specifically monitor the C. collinus marker molecules in emergency clinical and forensic cases. The simplicity and specificity make the ELISA superior to the other available techniques.


Production of polyclonal antibodies in ascitic fluid of mice: time and dose relationships. Further studies of the humoral immune response have been undertaken using ovine immunoglobulin G as the immunogen and BALB/C mice as the recipients. Mice immunized by the intraperitoneal route weekly for five weeks with an emulsion of the immunogen in Freund's complete adjuvant and given an intraperitoneal injection of pristane on day 14 produce relatively large volumes of ascitic fluid. A simple fluoroimmunoassay is used to determine specific antibody titres. The mice do not lose condition, remain active and eat normally throughout, although all show extensive granuloma formation within the peritoneal cavity. Tapping is performed before ascitic fluid production is excessive and each study is complete within 49 days, with a maximum of 5 taps. Only a single immunization site is required, since no higher antibody titres are obtained using combined intraperitoneal plus intramuscular plus subcutaneous immunization routes. A dose response study has shown that relatively large amounts (100 micrograms) of ovine immunoglobulin G must be injected to evoke a maximum response. Studies are in progress of various adjuvants and potential modulators of the immune response and of ways in which the immunogenicity of macromolecules can be changed. This approach can also be used to obtain antibodies for diagnostic purposes more rapidly than in rabbits and sheep.


Two-sites immunoradiometric assay using monoclonal antibodies for the determination of serum human sex hormone binding globulin. An immunoradiometric assay (IRMA) for the determination of human Sex Hormone Binding Globulin (SHBG) in serum is described. This IRMA uses three mouse monoclonal antibodies. Two monoclonals anti-human SHBG are coated on tubes and used as capture antibodies. The third monoclonal labeled with 125I completes the system, allowing the formation of the "sandwich". The detection limit of the assay is 2.5 femtomol SHBG per tube (250 pg/tube). Using this test for the measurement of SHBG and radioimmunoassays for the determination of total Testosterone and Estradiol, we calculated the Free Androgen Index (FAI) and the Free Testosterone. The results obtained were compared with the values of Free Testosterone measured by equilibrium dialysis. There is a close correlation between both calculated parameters and the levels of Free Testosterone, validating this SHBG assay.


Development of a high capacity radioimmunoassay procedure: ovine follicle stimulating hormone determination in blood plasma as a model. A sensitive, specific and accurate homologous radioimmunoassay (RIA) for ovine follicle stimulating hormone (oFSH) has been developed, using a [125I]oFSH tracer and a polyclonal rabbit anti-OFSH-serum at a final dilution of 1:224,000. The separation of free and antibody-bound tracer is based on the double antibody solid phase system. The assay was found to be specific for oFSH; cross-reactivity with oLH, oPrl and oGH was lower than 0.2%. The detection limit was 7 pg per tube. Inter- and intra- assay coefficients of variation (CV) were 3.7-8.5% and 6.6%, respectively. The use of a few selfmade appliances in combination with a well-considered time-schedule enabled the processing of three thousand blood plasma samples in triplicate within two weeks. The particular advantage of the method described here is the fast, easy and safe separation of free and antibody-bound tracer with minimal handling of radioactive tubes.


Detection of human anti-annexin autoantibodies by enzyme immunoassays. Annexins belong to a family of proteins characterized by calcium-dependent binding to the cytoskeleton and phospholipid surfaces. Basing on these properties annexins are discussed to be involved in the regulation of cytodynamic, anticoagulatory and antiinflammatory processes. Since autoantibodies against annexin I had been detected in patients suffering from inflammatory or autoimmune diseases, an impact on the pathophysiological outcome was assumed. Therefore we developed solid phase, enzyme-linked immunoassays for the quantitative determination of autoantibodies directed against six members of the annexin family. Some preliminary results obtained from sera of patients with malignant melanoma show a quite frequent presence of such autoantibodies. These data suggest that autoantibodies are generated against all annexins. Furthermore, in the individual patient autoantibodies of the IgG-type are monospecific, while about 1/4 of the IgM-type are directed against several annexins. These observations imply that for investigation of anti-annexin autoantibodies in inflammatory and autoimmune diseases as well as cancer all members of the annexin family have to be taken into consideration.


Rapid antibody capture assay for detection of group-A streptococci using monoclonal antibody and colloidal gold-monospecific polyvalent antibody conjugate. A rapid one step, sensitive and specific antibody capture assay for detection of group-A streptococci from the throat swabs of children is described. Monoclonal antibody either MA-106 or MA-107 specific for group-A streptococci polysaccharide (APS) was used as the capture antibody on nitrocellulose paper and rabbit monospecific polyvalent antibody conjugated with colloidal gold to detect the presence of antigen. The lower detection limit of this assay is 15.6ng APS/ml. The assay is specific for APS and failed to recognize polysaccharides obtained from group-B,-C,-G streptococci as well as Staphylococcus aureus. Antigen extracted from throat swabs of children who were positive for beta-hemolytic plaques (other than group-A streptococci) as seen on blood agar culture gave negative readings, thereby confirming the specificity of the assay for APS.


Improving the psychological status of children with asthma: a randomized controlled trial. Children with asthma are at special risk for problems in psychological functioning, as are children with other chronic illnesses. We conducted a controlled trial of a combined education and stress management program among children ages 6 to 14 years with asthma. Eighty-one children were randomly assigned to an intervention or a control group; 56 children completed data collection, 29 intervention and 27 control. Psychological status was assessed by the Child Behavior Checklist (CBCL) before and after the intervention, as were children's knowledge of asthma, stress (as measured by children's life events), and functional status (as indicated by such activities as school attendance, time playing with friends, and daily chore performance). Children in the intervention group had a significant improvement in the total Behavior Problems score (p less than .04) and Internalizing scale (p less than .01) on the CBCL and a significant increase in daily chores (p less than .04) compared with the control group. Before intervention, the two groups had statistically significant positive relationships between negative life events and behavior problems scores. After intervention, children in the control group still demonstrated a significant relationship between negative life events and total and Internalizing Behavior Problem scores, although participation in the intervention group negated that relationship. Children in the intervention group whose knowledge of asthma increased were more likely to report an increase in daily chores (p less than .02). We conclude that the intervention had a beneficial effect on psychological status and on children's daily activities. The effect may have occurred in part by decreasing the likelihood that perceived stress from negative life events led to poorer adjustment.


Home care for medically fragile children: impact on family health and well-being. The health status of 48 families providing home care for their medically fragile children was studied. Mothers, as the primary caregivers, experienced a greater decline in their physical health than did fathers or siblings. When the financial burden of providing care was greater and when the relationship with care providers was more strained, families had more physical illness symptoms. Who provided home care services for the medically fragile child influenced the psychosocial impact on the family. Care provided by home health aides was associated with greater negative impact, whereas care from professional nurses reduced the negative impact. The trend toward home care for medically fragile children has been accelerating; this study points to the importance of studying the impact on the family of this kind of care. Policy implications regarding the amount and quality of services and payment for them are discussed.


Impaired emotional health in children with mild reading disability. Children with reading disabilities are at risk for emotional difficulties. There is some evidence that reading-disabled children with high socioeconomic status may be at risk of having low self-esteem. We conducted a preliminary study of the impact of reading disability on children's self-esteem and emotional health in a select group of mildly impaired reading disabled children with well-educated parents who were well informed about reading disabilities. We interviewed 28 healthy, preadolescent boys and girls with mean reading delay of 9.0 months and 39 comparable nonreading disabled children who formed the control group. Children and parents completed the Harter's Self-Perception Profile. Parents completed the Achenbach Child Behavior Checklist and Rand Mental Health Survey. We found that these mildly impaired reading-disabled children were more anxious and less happy than were nondisabled students despite having well-informed, well-educated parents (mean level of education = 17.0 years). Reading-disabled children considered themselves to be less competent scholastically, and their parents rated them as less competent than nondisabled children on all measures of self-esteem. These well-educated mothers tended to underrate their child's perceived scholastic competence. Controlling for behavior problems, Achenbach Child Behavior Checklist scores for social competence were lower for reading-disabled children. These findings suggest that the mildly impaired reading-disabled children with high socioeconomic status are at risk for low self-esteem and poor emotional health.


Gender differences in infirmary use at a residential summer camp. Studies of health behavior in adults show that women report more morbidity and greater health service utilization than do men, despite lower mortality rates. Explanations involve social and biological gender differences in adult life. Infirmary utilization at a residential summer camp where parental influence is minimized was studied to determine whether these gender differences occur in the pediatric age group. Three hundred ninety-eight campers, 8 to 18 years old were studied. Girls were observed to make greater use of the infirmary than boys (p less than .01) and were especially likely to present with minor trauma, both musculoskeletal (p less than .05) and skin (p less than .01). No correlation was found between age and frequency of visits. Obvious morbidity was similar for boys and girls, in that no gender difference was observed in those visiting for definite medical indications alone or in those admitted. However, a difference was evident in those visiting for minor and trivial problems, particularly in those making three or more visits for minor and trivial problems (p less than .01). Although obvious morbidity was no different in campers, girls used the infirmary more than boys in a manner similar to that reported for adult health behavior. This suggests that gender differences occur earlier than suspected and are not simply related to adult social roles.


Personality development, problem solving, and behavior problems among preschool children with early histories of nonorganic failure-to-thrive: a controlled study. Using a prospective study design, 48 preschool children with early histories of nonorganic failure-to-thrive (NOFT) and 47 physically healthy children of comparable age, sex, birth order, and family demographics were compared on measures of problem solving and personality development (ego control and ego resiliency). In responding to structured measures, NOFT children demonstrated deficits in behavioral organization, ego control, and ego resiliency compared with controls. Parents also identified higher levels of behavioral symptoms in children with early histories of NOFT compared with controls. However, home observational measures of ego control and ego resiliency did not differentiate the two groups. Deficiencies in problem solving and personality development and increased levels of behavioral symptoms in preschool children with early histories of NOFT may affect their future socioemotional development and learning.


Psychometric differentiation of conduct disorder and attention deficit disorder with hyperactivity. Using a retrospective chart review procedure, children diagnosed as having attention deficit disorder with hyperactivity (ADDH), conduct disorder (CD), and ADDH+CD, as defined by DSM-III, were compared on measures of cognitive and academic functioning as well as on a continuous performance test (CPT). The groups did not differ in age or IQ, but children in the ADDH and ADDH+CD groups missed significantly more target stimuli on the CPT and performed significantly poorer on measures of academic achievement than did the pure CD group. However, poor CPT performance was not accounted for by learning problems. These findings in clinically referred patient groups are consistent with dimensional and epidemiological studies that suggest that ADDH may be a cognitively based disorder, whereas CD children are characterized by behavioral symptoms with fewer cognitive and learning deficits.


Decreased serum bicarbonate as a manifestation of undernutrition secondary to nonorganic failure-to-thrive. Eight of 101 children (8%) seen serially in consultation in an outpatient failure-to-thrive (FTT) clinic had isolated serum bicarbonate (TCO2) levels between 16 and 20 MEQ/dL (normal 22 to 30 MEQ/dL). None of these eight patients had signs or symptoms of renal disease, and all had nonorganic etiologic factors associated with their malnutrition. At the time of this review, follow-up TCO2 measurements were available for seven of the eight children who had attained normal weight (wt/age greater than fifth percentile on National Center for Health Statistics (NCHS) growth chart). All these children had normal TCO2 levels and no evidence of renal disease on follow-up more than 12 months later. Clinicians treating undernourished children who have low TCO2 measurements and have no signs or symptoms of renal disease and identified nonorganic factors, should consider a trial of therapies directed at nonorganic factors. Clinicians should be aware that a high percentage of undernourished children without other medical disease may have abnormal TCO2 measurements that correct after the institution of adequate caloric intake.


A complex chromosome rearrangement in a boy with autism. Autism is a rare behavioral phenotype defined by a qualitative impairment in reciprocal social interaction, impairment in communication and imaginative activity, and a markedly restricted repertoire of activities and interests. It is the most severe and prototypical form of the general category of Pervasive Developmental Disorders of Childhood. Using even strict diagnostic criteria, the currently described etiologies of autism are heterogeneous, with the majority of cases continuing to be idiopathic. At present, it is not clear whether autism is merely a behaviorally defined phenotype arising from diverse etiologies or a separate category of psychological dysfunction for which some unifying etiology exists. Complex chromosome rearrangements (CCR) are rare structural abnormalities involving at least three chromosomes and three or more break-points. We report a 6.5-year-old boy with classic infantile autism and a CCR involving chromosomes 1, 7, and 21. We discuss the possible relationship of his chromosome abnormality to the etiology of his autism.


A combined behavioral/pharmacological treatment of sleep-wake schedule disorder in Angelman syndrome. Angelman syndrome (AS) is a genetic disorder associated with a deletion on chromosome 15. Behavior problems among children with AS include sleep difficulties. Data are presented on the successful treatment of a sleep-wake schedule disorder (SWSD) in a 9-year-old boy with AS. The treatment program included behavioral and pharmacological components. During baseline, the child slept a mean of 1.9 hours per night and 1.3 hours during the day; night sleep was increased to a mean of 8.3 hours and day sleep was reduced to a mean of .08 hours after introduction of the full-treatment program. Medication was discontinued subsequently, and the child slept a mean of 7.8 hours during the night and .07 hours during the day. At 45-day follow-up, night sleep was maintained at 7.1 hours and day sleep remained stable at .29 hours. This is the first known report of an effective treatment of a SWSD in a child with AS.


Light and electron microscopic characterization of dopamine-immunoreactive axons in human cerebral cortex. The distribution and synaptic connections of dopamine axons were studied by light and electron microscopy in human cerebral cortex. For this purpose, dopamine immunoreactivity was characterized in apparently normal anteriolateral temporal cortex, which was removed to gain access to the medial temporal lobe during tumor excision or treatment of epilepsy. Nissl sections showed this to be granular neocortex. Dopamine fibers were distributed throughout this cortex, although there were relatively more fibers in layers I-II and in layers V-VIa, compared to layers III-IV and VIb, resulting in a bilaminar pattern of labeling. In all layers, fibers were seen to form numerous varicosities, and to vary in size from thick to very fine. Fibers were relatively straight, sparsely branched and were oriented in various planes within the cortex. However, in layer I, they often ran parallel to the pial surface. In order to analyze the functional interactions of dopamine fibers, individual cortical layers were surveyed for dopamine synapses. These were usually symmetrical (Gray's type II), although 13% of them were asymmetrical. Approximately 60% of dopamine synapses were made with dendritic spines, and 40% with dendritic shafts, and this ratio was similar in all layers. On both spines and shafts, it was common to see dopamine synapses closely apposed to an unlabeled asymmetric input, suggesting a dopamine modulation of excitatory input. Some postsynaptic dendritic shafts had features of pyramidal cells, including formation of spines. Since pyramidal cells are the major type of cortical spiny neuron, they probably represent the main target of dopamine synapses in this cortex. There were also dopamine profiles apposed to membrane densities on unlabeled axon terminals, suggesting another type of synaptic interaction. These findings provide the first documentation of dopamine synapses in the human cortex, and show that they form classical synaptic junctions. The location of these synapses on spines and distal dendrites, and their proximity to asymmetric synapses, suggest a modulatory role on excitatory input to pyramidal cells.


Changes in lamination and neuronal survival in the isthmo-optic nucleus following the intraocular injection of tetrodotoxin in chick embryos. We have studied how the development of the isthmo-optic nucleus (ION) is affected by electrical activity in the ION's axonal target territory, the contralateral retina. Electrical activity was blocked or reduced in the retina for various periods by tetrodotoxin injected intraocularly in different doses. The effects on the morphology of the retina appear to have been minor. During the ION's period of naturally occurring neuronal death (embryonic days 12 to 17), the injections substantially reduced this neuronal death and disrupted the development of lamination in the contralateral ION; there was also a lesser reduction in neuronal death in the ipsilateral ION. The dose of tetrodotoxin required to affect lamination was lower than that affecting neuronal death. Thus, the effects on neuronal death and on lamination were independent, since either could occur without the other. These effects were mediated by retrograde signals (probably two or more) from the eye; they occurred too early for the alternative anterograde route via the optic tectum (which projects to the ION) to be responsible. After embryonic day 17, the ION's response to intraocular tetrodotoxin changes abruptly from increased survival to total and rapid degeneration.


Maintenance of targeting errors by isthmo-optic axons following the intraocular injection of tetrodotoxin in chick embryos. We have studied the role of electrical activity in the elimination of axonal targeting errors, which is a normal process in brain development. The experiments were focused on the isthmo-optic nucleus (ION), which, in adults, projects in topographical order on the contralateral retina. During embryogenesis, however, a few isthmo-optic neurons project to the ipsilateral retina, and many project to topographically inappropriate parts of the contralateral one; both kinds of targeting error are known to be eliminated by the deaths of the parent neurons. We injected tetrodotoxin (TTX) intraocularly at embryonic days 13 and 15 and, on the latter, applied a retrograde label to the retina of the same eye. Embryos were fixed at embryonic day 17. In some embryos, the label was a peripherally placed fleck of the carbocyanine dye "diI"; the resulting retrogradely labeled neurons in the contralateral ION were much more widely scattered in the TTX-injected embryos than in controls (errors in topography). In other embryos, the label was a solution of rhodamine-B-isothiocyanate (RITC) injected into the vitreous body; this yielded several ipsilaterally labeled isthmo-optic neurons in the TTX-injected embryos, but virtually none in the controls. The numbers of both kinds of aberrantly projecting neuron approached those previously reported near the beginning of the ION's period of neuronal death. We conclude that electrical activity plays an important role in the elimination of axonal targeting errors in the chick embryo's isthmo-optic system.


Area 3a in the cat. II. Projections to the motor cortex and their relations to other corticocortical connections. It is well known that area 3a in the cat may monosynaptically influence the activity of neurons in the motor cortex. Much less information is available, however, on the anatomy of these connections. By using single or combined injections of different retrograde axonal tracers, we investigated the topography (horizontal and laminar) of area 3a neurons projecting to the motor cortex, and the anatomical relationships between these neurons and those projecting to other areas (2, 5, and SII) which, in turn, project to the motor cortex. Area 3a projects to all parts of area 4 gamma, to area 4 delta, and to the agranular area 6 in the lateral bank of the presylvian sulcus (area 6 alpha gamma), but not to other parts of areas 4 and 6. This projection exhibits a loose topographic organization along the mediolateral dimension of area 3a, and, in many cases, arises predominantly from the rostral half of this area. Although single small injections in the motor cortex produced two or more separate patches of retrograde labeling in 3a, after simultaneous injections of fluorochromes in two separate loci there often appeared in area 3a overlapping populations of neurons which were labeled retrogradely by each of the dyes, but with very few double-labeled neurons. In horseradish peroxidase (HRP) cases, 72% of area 3a neurons projecting to area 4 gamma were distributed in supragranular layers (mainly layer III), although the proportion of labeling in infragranular layers was larger when using fluorescent dyes. Double-labeled cells predominated in infragranular layers. These results have a bearing upon the functional roles that have been attributed to area 3a, as a cortical locus involved in muscle sensation, and a cortical relay to the motor cortex of rapid feedback information from muscle activity during movement.


Immunocytochemistry of dopamine in the brain of the locust Schistocerca gregaria. Catecholamine-induced histofluorescence studies have suggested a rich innervation of the locust brain by dopamine-containing neurons. To provide a basis for future studies on dopamine action in this insect, the location and morphology of neurons reacting with antisera against dopamine were investigated in the supraoesophageal ganglion of the locust, Schistocerca gregaria. In each brain hemisphere, about 100 interneurons in the midbrain and approximately 3,000 cells in the optic lobe show dopamine-like immunoreactivity. All major areas of the brain except the calyces of the mushroom body, the antennal lobe, large parts of the lobula, and some areas in the inferior lateral protocerebrum contain immunoreactive neuronal processes. The arborization patterns of most dopamine-immunoreactive cell types could be identified through detailed reconstructions. The central body exhibits the most intense immunostaining. It is innervated by at least 40 pairs of dopamine-immunoreactive neurons belonging to three different cell types. Additional arborizations of these neurons are in the superior protocerebrum and in the lateral accessory lobes. A group of 4 immunoreactive neurons with ramifications in the antennal mechanosensory and motor center gives rise to a dense meshwork of varicose fibers in the pedunculus and parts of the alpha- and beta-lobes of the mushroom body. Other cell types innervate the ventrolateral protocerebrum, the inferior protocerebrum and the posterior optic tubercles. Three descending neurons originating in the tritocerebrum exhibit dopamine-like immunoreactivity. In the optic lobe, about 3,000 columnar intrinsic neurons of the medulla and a group of centrifugal tangential cells with arborizations in the medulla and lamina are dopamine-immunoreactive.(ABSTRACT TRUNCATED AT 250 WORDS)


Diencephalic projections from the superficial and deep laminae of the medullary dorsal horn in the rat. An important function of the medullary dorsal horn (MDH) is the relay of nociceptive information from the face and mouth to higher centers of the central nervous system. We studied the central projection pattern of axons arising from the MDH by examining the axonal transport of Phaseolus vulgaris-leucoagglutinin (PHA-L). Labeled axon and axon terminal distributions arising from the MDH were analyzed at the light microscopic level. After large injections of PHA-L into both superficial and deep laminae of the MDH in the rat, labeled axons were observed in the nucleus submedius of the thalamus (SUB), ventroposterior thalamic nucleus medialis (VPM), ventroposterior thalamic nucleus parvicellularis (VPPC), posterior thalamic nuclei (PO), zona incerta (ZI), lateral hypothalamic nucleus (LH), and posterior hypothalamic nucleus (PH). Restriction of PHA-L into only the superficial laminae resulted in heavy axon and varicosity labeling in the SUB, VPM, PO, and VPPC and light labeling in LH. In contrast, after injections into deep laminae, labeled axons were mainly distributed in ZI and PH; some were also in VPM and LH, and fewer still in PO and SUB. Varicosities in VPM, SUB, and PO were significantly larger than those in VPPC, ZI, LH, and PH. Varicosity density was highest in SUB and lowest in the VPPC. We concluded that there are two distinct nociceptive pathways, one originating from the superficial MDH and terminating primarily in the dorsal diencephalon and the second originating from deep laminae of the MDH and terminating primarily in the ventral diencephalon. We propose that in the rat, input from the deeper laminae is primarily involved in the motivational-affective component of pain, whereas input from the superficial MDH is related to both the sensory-discriminative and motivational-affective component of pain.


Sex differences in the gross size of the rat neocortex. The pervasiveness of sex differences in the size of the rat cerebral cortex was investigated in ten littermate pairs of socially housed Long-Evans hooded rats at 90 days of age. Overall, the cortex was longer and wider in male than in female rats. Sex differences were detected in most cortical regions with this sample size; the exception was the temporal cortex. While some asymmetries were found, they did not vary with the sex of the animal nor were they consistently in one direction. The contribution of the cortical layers to sex differences in cortical thickness was examined in four locations: the primary motor cortex, the forelimb area of the sensorimotor cortex and the monocular and the binocular areas of the visual cortex. The layers that showed sex differences varied among the areas but were confined to layers II-III, V and VI. Sex differences in cortical size were widespread in the adult rat but showed some regional and laminar specificity.


Restoration of ascending noradrenergic projections by residual locus coeruleus neurons: compensatory response to neurotoxin-induced cell death in the adult rat brain. There is clinical and experimental evidence that monoamine neurons respond to lesions with a wide range of compensatory adaptations aimed at preserving their functional integrity. Neurotoxin-induced lesions are followed by increased synthesis and release of transmitter from residual monoamine fibers and by axonal sprouting. However, the fate of lesioned neurons after long survival periods remains largely unknown. Whether regenerative sprouting may contribute significantly to recovery of function following lesions which induce cell loss has been questioned. We have previously analyzed the response of locus coeruleus (LC) neurons to systemic administration of the noradrenergic (NE) neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) to adult rats. This drug causes ablation of nearly all LC axon terminals within 2 weeks after administration, followed by a profound loss of LC cell bodies 6 months later. The present study was conducted to determine the fate of surviving LC neurons and to characterize their potential for regenerative sprouting during a 16 month period after DSP-4 treatment. The time-course and extent of LC neuron degeneration were analyzed quantitatively in Nissl-stained sections, and the regenerative response of residual neurons was characterized by dopamine-beta-hydroxylase immunohistochemistry. The results document that LC neurons degenerate gradually after DSP-4 treatment, cell loss reaching on average 57% after 1 year. LC neurons which survive the lesion exhibit a vigorous regenerative response, even in those animals in which cell loss exceeds 60-70%. This regenerative process leads progressively to restoration of the NE innervation pattern in the forebrain, with some regions becoming markedly hyperinnervated. In stark contrast to the forebrain, very little reinnervation takes place in the brainstem, cerebellum and spinal cord. These findings suggest that regenerative sprouting of residual neurons is an important compensatory mechanism by which the LC may regain much of its functional integrity in the presence of extensive cell loss. Furthermore, regeneration of LC axons after DSP-4 treatment is region-specific, suggesting that the pattern of reinnervation is controlled by target areas. Elucidation of the factors underlying recovery of LC neurons after DSP-4 treatment may provide insights into the compensatory mechanisms of central neurons after injury and in disease states.


Sex differences in neuron number in the binocular area of the rat visual cortex. We have previously shown that the thickness of the binocular area of the primary visual cortex is sexually dimorphic in rats. In the present study, sex differences in the number of neurons in this cortical area were examined in nine littermate pairs of 90-day-old Long-Evans hooded rats. Cytoarchitectonic characteristics were used to define the binocular visual cortex, and its volume was estimated through three-dimensional reconstruction of serial coronal sections for each hemisphere. Neuronal and glial density as well as neuronal soma size were estimated from semithin sections through a stereological technique, the disector, in the same animals that were used to estimate volume. The volume of the binocular area was 19% greater in males than in females. While there were no sex differences in soma size or in neuronal density, the differences in the volume of the binocular area resulted in significant sex differences (male greater than female) in the number of neurons overall and in every layer, except layer IV. Glial density was not different between the sexes, but the total number of glial cells was higher in males than in females. These results demonstrate that the binocular visual cortex of the rat is sexually dimorphic in its volume and much of the difference is due to sex differences in the number of neurons and glial cells.


Localization of kynurenine aminotransferase immunoreactivity in the rat hippocampus. The localization and distribution of kynurenine aminotransferase (KAT), the biosynthetic enzyme of the excitatory amino acid receptor antagonist, kynurenic acid, was studied in the rat hippocampal formation with immunohistochemical methods. The enzyme was found mainly in glial cells that could be distinguished as 3 types on the basis of their shapes and locations. Typically, these cells shared the morphological features of astrocytes and exhibited glial fibrillary acidic protein immunoreactivity as demonstrated by a double-labeling technique. The distribution of KAT-containing glial cells was heterogeneous throughout the hippocampal formation. In the hippocampus, the stratum lacunosum-moleculare of Ammon's horn and the hilus contained a higher density of KAT-positive glial cells than other regions, whereas the lowest density of KAT glial cells was observed in the granule cell layer of the dentate gyrus and in the stratum radiatum of CA subfields. In the subicular complex, the density of KAT-containing glial cells was generally higher in the superficial than in the deep layer. Hippocampal neurons exhibiting KAT immunoreactivity, distinguished as nonpyramidal cells, were very few in number and mainly distributed in strata oriens and pyramidale of Ammon's horn. Substantially more KAT-positive neurons were observed in layers II and III of the subicular complex. The organization of cellular elements containing KAT may be of relevance for the function and possible dysfunction of kynurenic acid in the rat hippocampal formation.


Retrograde tracing of projections between the nucleus submedius, the ventrolateral orbital cortex, and the midbrain in the rat. The fluorescent tracers fluoro-gold and 1,1'-dioctadecyl-3,3,3,3-tetramethyl indocarbocyanine perchlorate were used as retrograde markers to examine reciprocal connections between the rat nucleus submedius and the ventrolateral orbital cortex. In addition, midbrain projections to each of these regions were examined. In the prefrontal cortex, we found that input from the nucleus submedius terminates rostrally within the lateral and ventral areas of the ventrolateral orbital cortex. Conversely, the cortical input to the nucleus submedius originates from the medial and dorsal parts of the ventrolateral orbital cortex. Our data also demonstrated that neurons from the ventrolateral periaqueductal gray and the raphe nuclei project to the midline nuclei of the thalamus, including a small projection to the nucleus submedius. We further determined that regions within the ventrolateral periaqueductal gray and raphe nuclei project to the ventrolateral orbital cortex, and that these regions overlap with those that project to the nucleus submedius. These findings suggest that the nucleus submedius might be part of a neural circuit involved in the activation of endogenous analgesia.


GABAergic neurons in the mammalian inferior olive and ventral medulla detected by glutamate decarboxylase immunocytochemistry. Neurons containing glutamic acid decarboxylase (GAD) (presumed GABAergic neurons) were mapped by immunocytochemistry in the ventral medulla of rat, rabbit, cat, rhesus monkey, and human, with emphasis on the inferior olive. In all species, three categories of GABAergic neurons were identified: periolivary neurons in the gray matter and the white matter surrounding the inferior olive, internuclear neurons located in the white matter between the subnuclei of the inferior olive, and intranuclear neurons located within the olivary gray matter. The intranuclear GABAergic neurons of the inferior olive had a characteristic morphology which differed from non-GABAergic olivary neurons; they were usually smaller, and, wherever their processes were stained, they had radiating, sparsely branching dendrites. They were also usually distinguished from the other GABAergic neurons by their smaller size. The intraolivary GABAergic neurons constituted only a minor proportion of the total olivary neuronal population, but they were concentrated in regions of the olive that varied by species. In the rat, they were situated in the rostral tip of the medial accessory olive and in the caudal subdivision of the dorsal accessory olive, while in the rabbit, they were located in the caudal two-thirds of the medial accessory olive, in the dorsal cap, and in the ventral lateral outgrowth. Such neurons were extremely rare in the cat; only a few were found in the rostral parts of the principal olive, the medial accessory olive, and the dorsal accessory olive. In the rhesus monkey, the principal olive and the lateral region of the rostral medial accessory olive contained most of the intranuclear GABAergic neurons, but some were also present in the dorsal accessory olive. In the human, such neurons occurred in the principal olive, the dorsal accessory olive and the rostral medial accessory olive, but as in the rhesus monkey, most were observed in the principal olive.


Common synaptic drive to segmentally homologous interneurons in the locust. The aim of the present study was to examine the pattern of synaptic interactions among a set of identified homologous interneurons in the segmental nervous system of the locust. This paper presents two main findings: first, serially homologous interneurons that are the progeny of neuroblast 7-4 in the mesothoracic, metathoracic, and first abdominal neuromeres of the locust central nervous system all receive synaptic drive from one and the same presynaptic interneuron. This interneuron has its entire arborization located in these three neuromeres of the central nervous system. It synapses with cells that are siblings, bilateral homologs, and serial homologs, and is itself connected monosynaptically with auditory afferents. The neuronal network that results comprises postsynaptic cells with the same developmental lineage. The second finding is that there is an additional set of synaptic connections among the homologous neurons themselves. All these connections are excitatory, and the pattern of information flow within the network is highly directional. This may relate to the morphologies of the neurons involved and will influence the contribution of homologs from different segments to behavior.


Development of high-affinity choline transport sites in rat forebrain: a quantitative autoradiography study with [3H]hemicholinium-3. The development of cholinergic terminals in rat brain has been quantitatively analyzed by [3H]hemicholinium-3 autoradiography. [3H]Hemicholinium-3 binds to high affinity choline transport sites, a specific marker for cholinergic neurons. In neonatal animals, kinetic and pharmacologic binding characteristics and regional distribution of [3H]hemicholinium-3 sites are consistent with specific cholinergic localization, as in the adult. The distribution of cholinergic terminals is described in the adult rat brain and during development, including heterogeneity of binding within several regions such as the striatum, nucleus accumbens, olfactory tubercle, cortex, and hippocampus. Early development and maturation vary greatly between brain regions. At embryonic day E18 and day 0, specific binding density is high only in the medial habenula. Development occurs primarily during the postnatal period in most brain regions examined. Many brain regions exhibit a lull in development between days 5 and 10, although the rate of development is highly region specific. Specific binding increases 2-12-fold between day 5 and adult animals, with adult density being achieved anywhere from day 15 to after day 21. The ontogeny of [3H]hemicholinium-3 binding sites generally occurs in a rostral to caudal direction. In the striatal body the characteristic lateral to medial gradient of binding site density is apparent by day 5, and development is more rapid in the lateral striatum. Patches of dense [3H]hemicholinium-3 binding coincident with acetylcholinesterase are observed on day 5 in the caudal striatum. The various patterns of cholinergic terminal development suggest that factors regulating cholinergic development are regional and complex.


Morphology of the dorsal cochlear nucleus in C57BL/6J and CBA/J mice across the life span. The morphology of the dorsal cochlear nucleus (DCN) was evaluated across the life span in inbred C57BL/6J (C57) and CBA/J (CBA) mice using 5 age groups (young adult to very old). C57 mice exhibit progressive cochlear sensorineural pathology and hearing loss during middle age; CBA mice have only modest sensorineural pathology late in life. DCN layers I, II, and III were evaluated histologically with serial sections stained for Nissl and fibers. DCN volume decreased with age in C57 mice, but the change began earliest and was most pronounced in layer III. In CBA mice, volume increased during the first year of life and decreased only in the oldest mice. All major DCN cell types were found in both strains at all ages. There was an age-related decrease in the mean size of neurons in C57 mice that was first observed in layer III. In CBA mice, only a nonsignificant trend toward smaller neurons was observed in the oldest mice. An age-related decline in the number of neurons in layer III (but not in layers I and II) occurred in C57 mice. Aged CBA mice exhibited no significant loss of DCN neurons. Thus, age-related changes in the DCN were much more pronounced in C57 mice than in CBA mice, and the changes in C57 mice were most pronounced in layer III. Because layer III receives most of the DCN's primary auditory input, it would be directly affected by age-related hearing loss and degeneration of spiral ganglion cells in C57 mice. This suggests that the age-related changes observed in DCN layer III of C57 mice are affected by progressive peripheral degenerative changes; when peripheral loss is minimal (CBA mice), less substantial age-related changes are observed.


Relationships between lipid availability and protein utilization during prolonged fasting. Mammals and birds adapt to prolonged fasting by mobilizing fat stores and minimizing protein loss. This strategy ends with an increase in protein utilization associated with behavioural changes promoting food foraging. Using the Zucker rat as a model, we have investigated the effect of severe obesity on this pattern of protein loss during long-term fasting. Two interactions between the initial adiposity and protein utilization were found. First, protein conservation was more effective in obese than in lean rats: fatty rats had a three times lower daily nitrogen excretion and proportion of energy expenditure deriving from proteins, and a lower daily protein loss in various muscles. This phase of protein sparing is moreover nine times longer in the fatty rats. Second, obese animals did not show the late increase in nitrogen excretion that occurred in their lean littermates. Total body protein loss during starvation was larger in fatty rats (57% versus 29%) and, accordingly, total protein loss was greater in their muscles. At the end of the experiment, lean and obese rats had lost 98% and 82%, respectively, of their initial lipid reserves, and fatty rats still had an obese body composition. These results support the hypothesis that in severely obese humans and animals a lethal cumulative protein loss is reached long before the exhaustion of fat stores, while the phase of protein conservation is still continuing. In contrast, in lean rats, survival of fasting seems to depend on the availability of lipid fuels.(ABSTRACT TRUNCATED AT 250 WORDS)


Adaptations of the reed frog Hyperolius viridiflavus (Amphibia: Anura: Hyperoliidae) to its arid environment. VI. The iridophores in the skin as radiation reflectors. Hyperolius viridiflavus possesses one complete layer of iridophores in the stratum spongiosum of its skin at about 8 days after metamorphosis. The high reflectance of this thin layer is almost certainly the result of multilayer interference reflection. In order to reflect a mean of about 35% of the incident radiation across a spectrum of 300-2900 nm only 30 layers of well-arranged crystals are required, resulting in a layer 10.5 microns thick. These theoretical values are in good agreement with the actual mean diameter of single iridophores (15.0 +/- 3.0 microns), the number of stacked platelets (40-100) and the measured reflectance of one complete layer of these cells (32.2 +/- 2.3%). Iridescence colours typical of multilayer interference reflectors were seen after severe dehydration. The skin colour turned from white (0-10% weight loss) through a copper-like iridescence (10-25% weight loss) to green iridescence (25-42%). In dry season state, H. viridiflavus needs a much higher reflectance to cope with the problems of high solar radiation load during long periods with severe dehydration stress. Dry-adapted skin contains about 4-6 layers of iridophores. The measured reflectance (up to 60% across the solar spectrum) of this thick layer (over 60 microns) is not in keeping with the results obtained by applying the multilayer interference theory. Light, scattered independently of wavelength from disordered crystals, superimposes on the multilayer-induced spectral reflectance. The initial parallel shift of the multilayer curves with increasing thickness and the almost constant ("white") reflectance of layers exceeding 60 microns clearly point to a changing physical basis with increasing layer thickness.


Effect of ambient temperature and E. coli endotoxin upon the plasma iron level in wild house mice in winter season. The effects of ambient temperatures of 10 degrees C and 30 degrees C and of E. coli endotoxin on brain temperature and plasma iron level were investigated in unrestrained wild house mice, Mus musculus. In control animals (i.p. saline-injected) exposed to cold environment the brain temperature decreased and plasma iron levels were lower than those observed under thermoneutral conditions (30 degrees C). Animals injected i.p. with endotoxin (0.5 micrograms.kg-1) and placed at 30 degrees C showed a drop in plasma iron level during the fever episode. The results provide strong evidence for a relationship between brain temperature and plasma iron level in control mice under thermoneutral conditions, and show that during cold exposure or after injection of endotoxin, there is no linear correlation between brain temperature and plasma iron. Moreover, it was found that cold stress influences plasma iron level and that this influence is not mediated by changes in brain temperature.


K+ current stimulation by Cl- in the midgut epithelium of tobacco hornworm (Manduca sexta). II. Analysis of Ba(2+)-induced K+ channel conduction noise. Chloride-stimulated K+ secretion by Manduca sexta midgut (5th-instar larvae) was measured as K(+)-carried short-circuit current of the tissue mounted in an Ussing chamber. "Microscopic" parameters, such as single-channel current and channel density for the rate-determining passive transport step across the basolateral goblet cell membrane (i.e. K+ channels), were estimated by means of current-fluctuation analysis of the K+ channel blockade by haemolymph-side Ba2+ ions. Ba2+ was equally effective with Cl- or gluconate (Glu-) as the principal ambient anion. The Ba(2+)-induced K+ channel conduction noise is reflected by a Lorentzian, or relaxation, noise component in the power spectrum of the K+ current fluctuations. A reduced Lorentzian plateau value, but an unchanged corner frequency, were observed when Cl- was replaced by Glu-. The results from the analysis of a "two-state" model of K+ channel block by Ba2+, with respect to the anion-replacement effects, suggest that the observed changes in K+ current and Lorentzian plateau value mirror a complex change of the underlying parameters: Cl- omission reduces single channel current but increases channel density so that the product of single channel current and channel density is smaller in Glu- than in Cl-. It seems likely that basolateral K+ channels (1) are subject to anionic gating ligands, and (2) depend on anions with respect to the rate of K+ transfer through an open K+ channel.


The role of elongation factors in protein synthesis rate variation in white teleost muscle. Protein synthesis-stimulating activity was assayed in the cytosolic fraction of white muscle from teleost fish (rainbow trout, carp) and of rat liver. In vitro protein synthesis-stimulating activity in the cytosolic fraction is reduced by food deprivation. The addition of elongation factors EF1, EF2, or EF1 + EF2 compensates for the starvation-induced loss of protein synthesis-stimulating activity in trout muscle cytosol. The action of EF2 is stronger than that of EF1 in this respect. However, EF1 enhances in vitro protein synthesis-stimulating activity in rat liver cytosol more than EF2. The EF2 concentration in the cytosolic fraction of white muscle from starved trout is significantly lower than in fed specimens.


Dietary protein levels affect growth and protein metabolism in trunk muscle of cod, Gadus morhua. Cod (Gadus morhua) of 50 g body weight were kept at 14 degrees C. The fish were fed ad libitum during 80 days a diet containing protein levels which in terms of total energy corresponded to 25%, 45% or 65%. Growth increased in accordance with protein-energy levels. The protein content per gram of wet weight of white trunk muscle was unchanged, as was the myofibrillar protein myosin heavy chain determined by the antigen-antibody reaction of the enzyme-linked immunosorbent assay. The amount of messenger ribonucleic acid (mRNA) coding for myosin heavy chain was lower at 25% than at 45% or 65% protein-energy intake, the differences being significant per gram of wet weight of muscle. Acid proteinase activity was highest at the lowest protein-energy intake. Glycogen content in muscle increased with the protein-energy levels. It is concluded that the metabolic response of white trunk muscle to graded protein-energy intake included a change in the capacity to synthesize myosin heavy chain as judged by its mRNA content. The protein content per gram of wet weight was unaffected by dietary protein-energy levels of 25%, 45% and 65%, but protein accretion and thus growth of the animals increased with the protein intake. Dietary protein-energy restriction caused a rise in acid proteinase activity and a decrease in content of mRNA for myosin heavy chain, resulting in a diminished growth rate at an unchanged protein content per gram of wet weight of muscle.


Renal, respiratory and ionic regulation in Atlantic salmon (Salmo salar L.) kelts following transfer from fresh water to seawater. Atlantic salmon may return to the sea after spawning in fresh water. These fish, known as kelts, reportedly show a limited ability to hypoosmoregulate. However, this study shows that fresh-water-adapted kelts exposed to seawater demonstrate rapid adaptation (within 48 h) in osmoregulatory parameters to values characteristics of seawater-adapted salmonids. The urine flow rate falls from 1.2 to 0.2 ml.kg-1.h-1 within 24 h. Over the same period, urine osmolality increases from 48 mosmol.kg-1 to become isosmotic with the plasma, and Mg2+ secretion by the kidney tubules elevates the urine concentration from 0.5 to 100 mmol.l-1. As is characteristic for marine teleosts, kelts drink seawater and process the ingested water in the gut to replace body water lost by osmosis to the hyperosmotic medium. Seawater exposure causes a marked hypoxia, arterial oxygen tension falling by 43% within minutes and persisting for at least 4 days at this low level. This is associated with large changes in blood pH and acid-base balance. The physiological mechanisms involved in adaptation to a hyperosmotic external medium are discussed, and the osmoregulatory capacity of kelts is compared with that of salmon at other stages of the life cycle.


Flight muscle catabolism during overnight fasting in a passerine bird, Eremophila alpestris. Brood-rearing passerine birds often have sparse lipid reserves coupled with potentially high energy demands. This may necessitate increased fasting protein catabolism; however, the largest source of protein, flight muscle, must be maintained. This problem was examined in the horned lark (Eremophila alpestris), a 28-g passerine. Overnight fasting caused significant depletion of protein in flight muscle and liver, but not in other muscle groups. Proteolytic enzyme activity of the flight muscle doubled during fasting. Biochemical and ultrastructural studies revealed that protein was depleted disproportionately from the sarcoplasm of flight muscle cells. Fasting caused a reduction in the protein-specific glycolytic capacity of flight muscle tissue. Oxidative capacity of the flight muscle, as measured by both in vivo and in vitro assays, was not significantly affected. The disproportionate catabolism of flight muscle sarcoplasmic protein may be due to a greater susceptibility to proteolysis, and not necessarily because it represents a source of redundant storage protein.


Role of behavior theory in behavioral medicine. Behavioral medicine is a multidisciplinary field that combines research methods and findings from behavioral and biomedical sciences. Many investigators in the field have tended to emphasize the contribution of the biomedical more than the behavioral sciences. This is evident in the emphasis on biological rather than behavioral outcomes and on the reductionist approach within the field to reduce mechanisms responsible for behavioral effects and disease to biological influences. There has been a similar shift in psychology toward mechanistic, bottom-up approaches to understanding mechanisms responsible for integrated and dynamic behavior. These shifts in emphasis have stimulated investigators to examine the use of biomedical methods and findings as causes and explanations for behavior, rather than to utilize newer findings in behavioral sciences. New advances in basic research on learning are used to illustrate that findings from behavioral science have implications for the field of contemporary behavioral medicine. Finally, the importance of developing new technologies for measuring behavior is presented.


Some structural changes that might facilitate the development of behavioral medicine. Three types of organizational change are considered that may facilitate the development of behavioral medicine teaching, practice, and research over the next decade. The first and most critical of these changes is a consideration of the role of a Department of Behavioral Medicine within the medical school. The second is support for the development of research collaborations across centers, with the advantage of bringing a broader array of expertise and facilities to the solution of health problems. Finally, some changes in the role and organization of scientific societies that support behavioral medicine are considered. It is argued that, without such changes, behavioral medicine will become overly narrow in its focus and will not have the impact on the medical arena that it might otherwise have.


Etiology and treatment of obesity: understanding a serious, prevalent, and refractory disorder. Obesity remains a leading public health problem because of its complications, prevalence, and resistance to change, despite record rates of dieting. Risk factors exist at both population and individual levels, thus obesity has diverse etiologies and consequences. Dieting is often grounded in the notion that the body can be molded at will and that the rewards justify the effort, creating a drive for unrealistic goals. This article argues for establishing a "reasonable weight," which may differ from health and aesthetic ideals. Intervention is less a matter of finding a "best" treatment but of finding the approach with "best fit" for the individual. Advances in treatment are most likely when research is driven by theory on the etiology of weight gain, the relapse process, and methods for permanent behavior change. This article integrates information on etiology, social beliefs about body weight, theory, and treatment into a comprehensive and compassionate model for intervention.


Smoking cessation: what have we learned over the past decade? This article discusses developments in understanding smoking and smoking cessation, methodological issues, and intervention approaches over the past 10 years. Although effective multisession clinic interventions have been developed, such programs reach relatively few smokers. This has led to self-help, work site, health care setting, and community interventions aimed at delivering less intensive programs to larger populations. Conceptual and empirical developments and trends within these above delivery contexts are reviewed, and avenues of research are identified. Nicotine replacement strategies have benefited from technological advances (e.g., transdermal patches) and present continuing challenges with respect to integration with behavioral strategies and incorporation into primary care medical settings. Research over the next decade should focus on the development of cost-effective interventions that can reach representative and high-risk smokers.


Behavioral and cognitive-behavioral approaches to chronic pain: recent advances and future directions. Behavioral and cognitive-behavioral approaches to chronic pain are receiving increasing attention from researchers and clinicians. This article reviews and highlights recent research advances and future research directions. Assessment research reviewed includes studies examining the social context of pain, the relationship of chronic pain to depression, cognitive variables affecting pain, and comprehensive assessment measures. Treatment outcome studies reviewed are those evaluating the effects of behavioral and cognitive-behavioral treatments for chronic pain. These studies focus on comparisons of behavioral treatment with control conditions, comparisons of two behavioral treatments, and prevention of chronic pain. Future directions for assessment and treatment research are outlined.


Psychological treatment of benign headache disorders. This article presents a selective summarization and critique of research on the psychological treatment of headache, with the primary focus on research appearing since 1980. Suggestions for future research directions and methodological improvements are included.


Psychoneuroimmunology: can psychological interventions modulate immunity? There is ample evidence from human and animal studies demonstrating the downward modulation of immune function concomitant with a variety of stressors. As a consequence, the possible enhancement of immune function by behavioral strategies has generated considerable interest. Researchers have used a number of diverse strategies to modulate immune function, including relaxation, hypnosis, exercise, classical conditioning, self-disclosure, exposure to a phobic stressor to enhance perceived coping self-efficacy, and cognitive-behavioral interventions, and these interventions have generally produced positive changes. Although it is not yet clear to what extent these positive immunological changes translate into any concrete improvements in relevant aspects of health, that is, alterations in the incidence, severity, or duration of infectious or malignant disease, the preliminary evidence is promising.


Psychological interventions for cancer patients to enhance the quality of life. Although the thrust of the nation's cancer objectives for the year 2000 is prevention and screening, each year approximately 1 million Americans are diagnosed and must cope with the disease and treatments. They do so with the aid of family, friends, and the health care system, but accumulating data suggest that psychological interventions may be important for reducing emotional distress, enhancing coping, and improving "adjustment." Experimental and quasi-experimental studies of psychological interventions are reviewed, and discussion of treatment components and mechanism is offered. A final section discusses future research directions and challenges to scientific advance.