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Hormone coexpression in the adult toad endocrine pancreas: double-label immunofluorescence under basal conditions and after glucose injection.

We have investigated the type and frequency of hormone coexpression in the endocrine pancreas of amphibians both under basal conditions and after sustained glucose loading. Adult male specimens of the wild toad Bufo arenarum were injected with a 50% (w/v) glucose solution (2 g/100 g) for 2 days, while control animals received an equal volume of the vehicle. Serum glucose levels were measured at the time of sacrifice and the pancreatic free lobe was processed for light microscopy. A double-labeling immunofluorescence study was performed for the detection of insulin (I), glucagon (G), somatostatin (S), and pancreatic polypeptide (PP). Heterospecific antisera against the following hormone combinations were used for their detection and immunocytochemical localization: I+G, I+PP, G+PP, S+G, and S+PP; visualization of the reacted IgG's was effected by fluorescein- and rhodamine-conjugated species-specific antibodies as fluorophores. Intracellular hormone coexpression was found to occur in the combinations G+PP, S+G, and S+PP. Moreover, glucose administration caused, together with a marked hyperglycemia (123 +/- 17 vs 23 +/- 1 mg/dl; P < 0.05), a decrease in the fraction of cells containing both G and PP together (from 106.3 +/- 8.1 to 26 +/- 4 cell/mm2) along with a reciprocal rise in the number of cells possessing G alone (from 128.7-152.3 to 235.9-274 cell/mm2). The fewer number of cells coexpressing either of the other two hormone combinations, however, were unaffected by glucose injection. With respect to the simultaneous measurement of I+G and I+PP, no cells were detected with both hormones of either pair, and the I-containing cells were more frequent in each instance in the control toads (264.8 +/- 22.3 to 269.2 +/- 27 cell/mm2). For both combinations, however, this value diminished significantly in the glucose-treated animals (108 +/- 2 cell/mm2 for I+G and 112.1 +/- 7. 8 cell/mm2 for I+PP). While the G-containing cells became more numerous (rising to 235.9 +/- 12.4, 274 +/- 26, and 250.4 +/- 23.7 cell/mm2 for I-G, G-PP, and G-S combinations, respectively), the PP- and S-containing cells remained unaffected. We conclude that the copresence of different hormones within the same cell is a relatively common finding in the non-I-secreting elements of the adult toad pancreas and that the proportions of specific cell types are affected by glucose administration. We thus propose that intracellular hormonal coexpression in this fashion may well represent a rapid and efficient regulatory mechanism for compensating for the metabolic stress imposed by glucose loading.