Summary
The aim of this study was to localize cells immunoreactive for glutamate decarboxylase (GAD), the enzyme of GABA synthesis, in pyloric and oxyntic regions of the rat stomach as well as in the rat and mouse pancreas. GAD immunocytochemistry was carried out on polyethylene glycol or cryostat sections of alkaline paraformaldehyde fixed tissue, with simultaneous immunolabelling of various gastro-pancreatic hormones for topographical comparison. In the rat stomach, nerve fibers displaying intense GAD-like immunoreactivity were seen in the myenteric plexus, the circular muscular layer, the submucosa and the lamina propria of the mucosa. But, they were absent from the submucous plexus. Colchicine treatment of the rats allowed to detect some labelled perikarya in the myenteric plexus suggesting that the GABAergic innervation is at least partly intrinsic to the stomach. In the oxyntic and pyloric mucosa, endocrine cells appeared immunostained for GAD. However, the nature of their hormones remained unknown since double immunodetections revealed that they were immunoreactive neither for gastrin nor for somatostatin. In the rat and mouse pancreas, GAD-like immunoreactivity was found in islet cells which corresponded only to insulin-secreting cells. Somatostatin-, glucagon- and pancreatic polypeptide-immunopositive cells were devoid of GAD immunolabelling. No GAD-like immunoreactivity was detected in the exocrine tissue and innervation. These results strenghten the hypothesis that GABA is not only a neurotransmitter in the stomach but that it could also be an endocrine or paracrine factor in the stomach and pancreas.
Similar content being viewed by others
References
Baekkeskov S, Aanstoot HJ, Christgau S, Reetz A, Solimena M, Cascalho M, Folli F, Richter-Olesen H, De Camilli P (1990) Identification of the 64K autoantigen in insulin-dependent diabetes as the GABA-synthesizing enzyme glutamic acid decarboxylase. Nature 347:151–156
Berod A, Hartman BK, Pujol JF (1981) Importance of fixation in immunohistochemistry: use of formaldehyde solutions at variable pH for the localization of tyrosine hydroxylase. J Histochem Cytochem 29:844–850
Davanger S, Ottersen OP, Storm-Mathisen J (1987) Immunocytochemical localization of GABA in cat myenteric plexus. Neurosci Lett 73:27–32
Davanger S, Ottersen OP, Storm-Mathisen J (1989) GABA-immunoreactive cells in the rat gastrointestinal epithelium. Anat Embryol 179:221–226
Erdö SL, Wolff JR (1988) Releasable, non-neuronal GABA pool in rat stomach. Eur J Pharmacol 156:165–168
Erdö SL, Ezer E, Matuz J, Wolff JR, Amenta F (1989) GABAA receptors in the rat stomach may mediate mucoprotective effects. Eur J Pharmacol 165:79–86
Erdö SL, De Vincentis G, Amenta F (1990) Autoradiographic localization of [3H]muscimol binding sites in rat stomach: evidence for mucosal GABAA receptors. Eur J Pharmacol 175:351–354
Gamrani H, Harandi M, Belin MF, Dubois MP, Calas A (1984) Direct electron microscopic evidence for the coexistence of GABA uptake and endogenous serotonin in the same rat central neurons by coupled radioautographic and immunocytochemical procedures. Neurosci Lett 48:25–30
Garry DJ, Sorenson RL, Coulter HD (1987a) Ultrastructural localization of gamma amino butyric acid immunoreactivity in B cells of the rat pancreas. Diabetologia 30:115–119
Garry DJ, Coulter HD, MacIntee TJ, Wu JY, Sorenson RL (1987b) Immunoreactive GABA transaminase within the pancreatic islet is localized in mitochondria of the B-cell. J Histochem Cytochem 35:831–836
Garry DJ, Garry MG, Sorenson RL (1988) Ultrastructural immunocytochemical localization of l-glutamate decarboxylase and GABA in rat pancreatic zymogen granules. Cell Tissue Res 252:191–197
Gilon P (1990) Le GABA et les cellules endocrines du système gastro-entéro-pancréatique. Thesis, University of Louvain, Louvain-la-Neuve
Gilon P, Remacle C, Janssens de Varebeke P, Pauwels G, Hoet JJ (1987a) GABA content and localization of high-affinity GABA uptake during the development of the rat pancreas. Cell Mol Biol 33:573–585
Gilon P, Reusens-Billen B, Remacle C, Janssens de Varebeke P, Pauwels G, Hoet JJ (1987b) Localization of high-affinity GABA uptake and GABA content in the rat duodenum during development. Cell Tissue Res 249:593–600
Gilon P, Campistron G, Geffard M, Remacle C (1988) Immunocytochemical localization of GABA in endocrine cells of the rat entero-pancreatic system. Biol Cell 62:265–273
Gilon P, Remacle C (1989) High-affinity GABA uptake in a subpopulation of somatostatin cells in rat pancreas. J Histochem Cytochem 37:1133–1139
Gilon P, Mallefet J, De Vriendt C, Pauwels S, Geffard M, Campistron G, Remacle C (1990) Immunocytochemical and autoradiographic studies of the endocrine cells interacting with GABA in the rat stomach. Histochemistry 93:645–654
Gottlieb DI, Chang YC, Schwob JE (1986) Monoclonal antibodies to glutamic acid decarboxylase. Proc Natl Acad Sci USA 83:8808–8812
Guo YS, Thompson JC, Singh P (1989) Effect of γ-aminobutyric acid on bombesin-evoked release of somatostatin and gastrin from isolated rat stomach. Regul Pep 24:179–186
Hakanson R, Alumets J, Stewart JA, Sundler F (1978) Does glutamate decarboxylase occur in endocrine cells of gut and pancreas? Scand J Gastroenterol 13 [Suppl 49]:74
Harty RF, Franklin PA (1983) GABA affects the release of gastrin and somatostatin from rat antral mucosa. Nature 303:623–624
Harty RF, Franklin PA (1986) Cholinergic mediation of γ-aminobutyric acid-induced gastrin and somatostatin release from rat antrum. Gastroenterology 91:1221–1226
Harty RF, Murthy SNS (1986) GABAergic mechanisms in the endocrine stomach and duodenum: their possible functional significance. In: Erdö SL, Bowery NG (eds) GABAergic mechanisms in the mammalian periphery. Raven Press, New York, pp 339–352
Harty RF, Carr T, Boharski MG, Lassiter DC, Egan PT (1986) Cholinergic and γ-aminobutyric acid neurotransmitter release from antral mucosal and submucosal neurons. Can J Physiol Pharmacol (Suppl) 64:39
Hills JM, Jessen KR, Mirsky R (1987) An immunohistochemical study of the distribution of enteric GABA-containing neurons in the rat and guinea-pig intestine. Neuroscience 22:301–312
Hougaard Dm, Nielsen JH, Larsson LI (1986) Localization and biosynthesis of polyamines in insulin-producing cells. Biochem J 238:43–47
Jesen KR, Mirsky R, Hills JM (1987) GABA as an autonomic neurotransmitter: studies on intrinsic GABAergic neurons in the myenteric plexus of the gut. TINS 10:255–262
Jessen KR, Hills JM, Limbrick AR (1988) GABA immunoreactivity and 3H-GABA uptake in mucosal epithelial cells of the rat stomach. Gut 29:1549–1556
Klosen P, van den Bosch de Aguilar P (1990) Polyethylene-glycol sections for immunocytochemistry of neural antigens: a reappraisal. Eur J Neurosci [Suppl] 3:249
Koop H, Arnold R (1986) Control of rat gastric somatostatin release by γ-aminobutyric acid (GABA). Horm Metab Res 18:94–97
Krantis A, Webb T (1989) Autoradiographic localization of [3H] γ-aminobutyric acid in neuronal elements of the rat gastric antrum and intestine. J Auton Nerv Sys 29:41–48
Lechago J (1987) The endocrine cells of the digestive tract. General concepts and historic perspective. Am J Surg Pathol 11 [Suppl 1]:63–70
Legay F, Pelhate S, Tappaz ML (1986) Phylogenesis of brain glutamic acid decarboxylase from vertebrates: immunochemical studies. J Neurochem 46:1478–1486
Legay F, Henry S, Tappaz ML (1987a) Monoclonal antibodies against rat brain glutamic acid decarboxylase (GAD). Neurochem Int 10:287–294
Legay F, Henry S, Tappaz ML (1987b) Evidence for two distinct forms of native glutamic acid decarboxylase in rat brain soluble extract: an immunoblotting study. J Neurochem 48:1022–1026
Li CY, Ziesmer SC, Lazcano-Villareal O (1987) Use of azide and hydrogen peroxide as an inhibitor for endogenous peroxidase in the immunoperoxidase method. J Histochem Cytochem 35:1457–1460
Neale EA, Oertel WH, Bowers LM, Weise VK (1983) Glutamate decarboxylase immunoreactivity and γ-(3H)aminobutyric acid accumulation within the same neurons in dissociated cell cultures of cerebral cortex. J Neurosci 3:376–382
Oertel WH, Schmechel DE, Tappaz ML, Kopin IJ (1981a) Production of a specific antiserum to rat brain glutamic acid decarboxylase by injection of an antigen-antibody complex. Neuroscience 6:2689–2700
Oertel WH, Schmechel DE, Mugnaini E, Tappaz ML, Kopin IJ (1981b) Immunocytochemical localization of glutamate decarboxylase in rat cerebellum with a new antiserum. Neuroscience 6:2715–2735
Okada Y (1979) Properties of glutamate decarboxylase (GAD) in the human insulinoma, a non-neural tissue. Proc Jpn Acad 55:514–517
Okada Y (1986) Localization and function of GABA in the pancreatic islets. In: Erdö SL, Bowery NG (eds) GABAergic mechanisms in the mammalian periphery. Raven Press, New York, pp 223–240
Okada Y, Taniguchi H, Baba S (1982) High concentration of GABA in the pancreatic islets with special emphasis on B cells. In: Okada Y, Roberts E (eds) Problems in GABA research from brain to bacteria. Excerpta Medica, Amsterdam, pp 379–386
Rahier J, Pauwels S, Dockray GJ (1987) Biosynthesis of gastrin. Localization of the precursor and peptide products using electron microscopic-immunogold methods. Gastroenterology 92:1146–1152
Reisert I, Wöhrle M, Pilgrim C (1985) Quantitative assessment of GABA-uptake sites in the neural lobe by electron-microscopic autoradiography. Cell Tissue Res 241:581–584
Rorsman P, Berggren PO, Bokvist K, Ericson H, Möhler H, Östenson CG, Smith PA (1989) Glucose-inhibition of glucagon secretion involves activation of GABAA-receptor chloride channels. Nature 341:233–236
Sakanaka M, Shibasaki T, Lederis K (1987) Improved fixation and cobalt-glucose oxidase-diaminobenzidine intensification for immunohistochemical demonstration of corticotropin-releasing factor in rat brain. J Histochem Cytochem 35:207–212
Sakaue M, Saito N, Tanaka C (1987) Immunohistochemical localization of gamma-aminobutyric acid (GABA) in the rat pancreas. Histochemistry 86:365–369
Solcia E, Usellini L, Buffa R, Rindi G, Villani L, Zampatti C, Silini E (1987) Endocrine cells producing regulatory peptides. Experientia 43:839–850
Staines WA, Meister B, Melander T, Nagy JI, Hökfelt T (1988) Three-color immunofluorescence histochemistry allowing triple labeling within a single section. J Histochem Cytochem 36:145–151
Sternberger LA, Joseph SA (1979) The unlabeled antibody method. Contrasting color staining of paired pituitary hormones without antibody removal. J Histochem Cytochem 27:1424–1429
Taniguchi H, Murakami K, Yoshioka M, Ejiri K, Ishihara K, Baba S, Okada Y (1982a) GABA and insulin in pancreatic islets. In: Okada Y, Roberts E (eds) Problems in GABA research from brain to bacteria. Excerpta Medica, Amsterdam, pp 387–405
Taniguchi H, Yoshioka M, Ejiri K, Ishihara K, Tamagawa M, Hirose Y, Ishihara K, Utsumi M, Baba S, Okada Y (1982b) Suppression of somatostatin release and increase of somatostatin content in pancreatic islets by GABA. In: Okada Y, Roberts E (eds) Problems in GABA research from brain to bacteria. Excerpta Medica, Amsterdam, pp 406–412
Tappaz ML, Wassef M, Oertel WH, Paut L, Pujol JF (1983) Light- and electronmicroscopic immunocytochemistry of glutamic acid decarboxylase (GAD) in the basal hypothalamus: morphological evidence for neuroendocrine γ-aminobutyrate (GABA). Neuroscience 9:271–287
Thirbly RC, Stevens MH, Blair AJ, Petty F, Crawford IL, Taylor IL, Walsh JH, Feldman M (1988) Effect of GABA on basal and vagally mediated gastric acid secretion and hormone release in dogs. Am J Physiol 254:G723-G731
Vincent SR, Hökfelt T, Wu JY, Elde RP, Morgan LM, Kimmel JR (1983) Immunohistochemical studies of the GABA system in the pancreas. Neuroendocrinology 36:197–204
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Gilon, P., Tappaz, M. & Remacle, C. Localization of GAD-like immunoreactivity in the pancreas and stomach of the rat and mouse. Histochemistry 96, 355–365 (1991). https://doi.org/10.1007/BF00271357
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00271357