Upregulation of alpha-synuclein expression in the rat cerebellum in experimental hepatic encephalopathy.

AIMS: The overexpression of alpha-synuclein has been associated with neurodegenerative diseases, especially when the protein aggregates to form insoluble structures. The present study examined the effect of chronic hyperammonaemia on alpha-synuclein expression in the rat cerebellum following portacaval anastomosis (PCA). METHODS: Immunohistochemical and western blot determinations were performed 1 month and 6 months after the PCA procedure. RESULTS: A time-dependent increase in alpha-synuclein expression was seen in the cerebellar grey matter compared with the controls. At 1 month post PCA, alpha-synuclein-immunopositive material was observed in the molecular layer, while the Purkinje cells showed weak alpha-synuclein expression, and alpha-synuclein aggregates were observed throughout the granular layer. At 6 months post PCA, alpha-synuclein expression was significantly increased compared with the controls. alpha-synuclein-immunostained astroglial cells were also found; the Bergmann glial cells showed alpha-synuclein-positive processes in the molecular layer of PCA-exposed rats, and in the granular layer, perivascular astrocytes showed intense alpha-synuclein immunoreactivity, as indicated by colocalization of alpha-synuclein with glial fibrillary acidic protein (GFAP). In addition, ubiquitin-immunoreactive inclusions were present in PCA-exposed rats, although they did not colocalize with alpha-synuclein. Western blotting performed at 6 months post PCA showed a reduction in the level of soluble alpha-synuclein compared with 1 month post PCA and the controls; this reduction was concomitant with an increase in the insoluble form of alpha-synuclein. CONCLUSIONS: Although the precise mechanism by which alpha-synuclein aggregates in PCA-treated rats remains unknown, the present data suggest an important role for this protein in the onset and progression of hepatic encephalopathy, probably via its expression in astroglial cells.

Microvesicles: ROS scavengers and ROS producers.

This review analyzes the relationship between microvesicles and reactive oxygen species (ROS). This relationship is bidirectional; on the one hand, the number and content of microvesicles produced by the cells are affected by oxidative stress conditions; on the other hand, microvesicles can directly and/or indirectly modify the ROS content in the extra- as well as the intracellular compartments. In this regard, microvesicles contain a pro-oxidant or antioxidant machinery that may produce or scavenge ROS: direct effect. This mechanism is especially suitable for eliminating ROS in the extracellular compartment. Endothelial microvesicles, in particular, contain a specific and well-developed antioxidant machinery. On the other hand, the molecules included in microvesicles can modify (activate or inhibit) ROS metabolism in their target cells: indirect effect. This can be achieved by the incorporation into the cells of ROS metabolic enzymes included in the microvesicles, or by the regulation of signaling pathways involved in ROS metabolism. Proteins, as well as miRNAs, are involved in this last effect.

Hepatocyte growth factor activates several transduction pathways in rat pancreatic acini.

The receptor of hepatocyte growth factor (HGF), c-met induces different physiological responses in several cell types. Little is known about the role of HGF in exocrine pancreas. However, abnormal HGF signaling has been strongly implicated in pancreatic tumorigenesis and association of HGF with pancreatitis has been demonstrated. We have studied the presence of c-met and activation of their intracellular pathways associated in rat pancreatic acini in comparison with cholecystokinin (CCK) and epidermal growth factor (EGF). C-met expression in rat exocrine pancreas was identified by immunohistochemistry and immunoprecipitation followed by Western analysis. Tyrosine phosphorylation of c-met is strongly stimulated as well as kinase pathways leading to ERK1/2 cascade. HGF, but not CCK or EGF, selectively caused a consistent increase in the amount of p85 regulatory subunit of PI3-K present in anti-phosphotyrosine immunoprecipitates. Downstream of PI3-K, HGF increased Ser473 phosphorylation of Akt selectively, as CCK or EGF did not affect it. HGF selectively stimulated tyrosine phosphorylation of phosphatase PTP1D. HGF failed to promote the well-known CCK effects in pancreatic acini such as amylase secretion and intracellular calcium mobilization. Although HGF shares activation of ERK1/2 with CCK, we demonstrate that it promotes the selective activation of intracellular pathways not regulated by CCK or EGF. Our results suggest that HGF is an in vivo stimulus of pancreatic acini and provide novel insight into the transduction pathways and effects of c-met/HGF in normal pancreatic acinar cells.

Neuronal and inducible nitric oxide synthase expression in the rat cerebellum following portacaval anastomosis.

In order to determine the role of neuronal nitric oxide synthase (nNOS) and inducible nitric oxide synthase (iNOS) in the pathogenesis of experimental hepatic encephalopathy (HE), the expression of both was analyzed in the cerebellum of rats 1 month and 6 months after performing portacaval anastomosis (PCA). In control cerebella, nNOS immunoreactivity was mainly observed in the molecular layer (ML), whereas the Purkinje cells did not express nNOS. However, nNOS expression was detected in the Purkinje cells at 1 month after PCA, correlating with a decrease in nNOS expression in the ML--part of an overall reduction in cerebellar nNOS concentrations (as determined by Western blotting). At 6 months post-PCA, a significant increase in nNOS expression was observed in the ML, as well as increased nNOS immunoreactivity in the Purkinje cells. nNOS immunoreactivity was also observed in the Bergmann glial cells of PCA-treated rats. While no immunoreactivity for iNOS was seen in the cerebella of control rats, iNOS immunoreactivity was significantly induced in the cerebellum 1 month after PCA. In addition, the expression of iNOS was greater at 6 months than at 1 month post-PCA. Immunohistochemical analysis revealed this iNOS to be localized in the Purkinje cells and Bergmann glial cells. The induction of iNOS in astroglial cells has been associated with pathological conditions. Therefore, the iNOS expression observed in the Bergmann glial cells might play a role in the pathogenesis of HE, the harmful effects of PCA being caused by them via the production of excess nitric oxide. These results show that nNOS and iNOS are produced in the Purkinje cells and Bergmann glial cells following PCA, implicating nitric oxide in the pathology of HE.

Identification of a protein-tyrosine phosphatase (SHP1) different from that associated with acid phosphatase in rat prostate.

Using [32P]poly(Glu,Tyr) as substrate, we have identified, for the first time, in the rat prostatic gland a protein-tyrosine phosphatase activity different from that associated with prostatic acid phosphatase. Concanavalin A-Sepharose 4B was used to separate the two protein-tyrosyl phosphatases activities. The activity retained by the lectin had characteristics of the prostatic acid phosphatase. It was sensitive to inhibition by PNPP and the optimum pH shifted towards physiological values when [32P]poly(Glu,Tyr) was used as substrate. However, the major protein-tyrosine phosphatase activity was not retained by the lectin, and corresponded, at least in part, to SHP1 as probed by the presence of the protein, its mRNA and the loss of PTPase activity after immunodepletion of SHP1. This enzyme is localized within the epithelial cells. Thus, the coexistence of two protein-tyrosine phosphatase activities in rat prostate, one associated with the acid phosphatase and the other related to SHP1, makes it necessary to analyze the importance of both activities in vivo and their possible function regarding prostatic cell growth and its regulation.

Redistribution of protein kinase C isoforms in rat pancreatic acini during lactation and weaning.

Freshly enzymatically isolated pancreatic acini from lactating and weaning Wistar rats were used to investigate the role of protein kinase C (PKC) isoforms during these physiologically relevant pancreatic secretory and growth processes. The combination of immunoblot and immunohistochemical analysis shows that the PKC isoforms alpha, delta, and epsilon are present in pancreatic acini from control, lactating and weaning rats. A vesicular distribution of PKC-alpha, -delta, and -epsilon was detected by immunohistochemical analysis in the pancreatic acini from all the experimental groups. PKC-delta showed the strongest PKC immunoreactivity (PKC-IR). In this vesicular distribution, PKC-IR was located at the apical region of the acinar cells. No differences were observed between control, lactating and weaning rats. However, the immunoblot analysis of pancreatic PKC isoforms during lactation and weaning showed a significant translocation of PKC-delta from the cytosol to the membrane fraction when compared with control animals. Translocation of PKC isoforms (alpha, delta and epsilon) in response to 12-O-tetradecanoyl phorbol 13-acetate (TPA) 1 microM (15 min, 37 degrees C) was comparable in pancreatic acini from control, lactating and weaning rats. In the control group, a significant translocation of all the isoforms (alpha, delta and epsilon) from the cytosol to the membrane was observed. The PKC isoform most translocated by TPA was PKC-delta. In contrast, no statistically significant increase in PKC-delta translocation was detected in pancreatic acini isolated from lactating or weaning rats. These results suggest that the PKC isoforms are already translocated to the surface of the acinar cells from lactating or weaning rats. In addition, they suggest that isoform specific spatial PKC distribution and translocation occur in association with the growth response previously described in the rat exocrine pancreas during lactation and weaning.

Glutamine synthetase in brain: effect of ammonia.

Glutamine synthetase (GS) in brain is located mainly in astrocytes. One of the primary roles of astrocytes is to protect neurons against excitotoxicity by taking up excess ammonia and glutamate and converting it into glutamine via the enzyme GS. Changes in GS expression may reflect changes in astroglial function, which can affect neuronal functions. Hyperammonemia is an important factor responsible of hepatic encephalopathy (HE) and causes astroglial swelling. Hyperammonemia can be experimentally induced and an adaptive astroglial response to high levels of ammonia and glutamate seems to occur in long-term studies. In hyperammonemic states, astroglial cells can experience morphological changes that may alter different astrocyte functions, such as protein synthesis or neurotransmitters uptake. One of the observed changes is the increase in the GS expression in astrocytes located in glutamatergic areas. The induction of GS expression in these specific areas would balance the increased ammonia and glutamate uptake and protect against neuronal degeneration, whereas, decrease of GS expression in non-glutamatergic areas could disrupt the neuron-glial metabolic interactions as a consequence of hyperammonemia. Induction of GS has been described in astrocytes in response to the action of glutamate on active glutamate receptors. The over-stimulation of glutamate receptors may also favour nitric oxide (NO) formation by activation of NO synthase (NOS), and NO has been implicated in the pathogenesis of several CNS diseases. Hyperammonemia could induce the formation of inducible NOS in astroglial cells, with the consequent NO formation, deactivation of GS and dawn-regulation of glutamate uptake. However, in glutamatergic areas, the distribution of both glial glutamate receptors and glial glutamate transporters parallels the GS location, suggesting a functional coupling between glutamate uptake and degradation by glutamate transporters and GS to attenuate brain injury in these areas. In hyperammonemia, the astroglial cells located in proximity to blood-vessels in glutamatergic areas show increased GS protein content in their perivascular processes. Since ammonia freely crosses the blood-brain barrier (BBB) and astrocytes are responsible for maintaining the BBB, the presence of GS in the perivascular processes could produce a rapid glutamine synthesis to be released into blood. It could, therefore, prevent the entry of high amounts of ammonia from circulation to attenuate neurotoxicity. The changes in the distribution of this critical enzyme suggests that the glutamate-glutamine cycle may be differentially impaired in hyperammonemic states.

Response of rat cerebral somatostatinergic system to a high ammonia diet.

It has been reported that ingestion of an ammonium-containing diet produces hyperammonemia without encephalopathy, thus permitting the study of the specific effects of ammonia toxicity. The present study investigated the rat cerebral somatostatinergic system using this experimental model of hyperammonemia. Wistar rats were fed a high ammonia diet prepared by mixing a standard diet with ammonium acetate (20% w/w); in addition, 5 mM of ammonium acetate was added to their water supply. Control rats were fed with a standard diet. The animals were sacrificed at 3, 7 and 15 days of ammonia ingestion. Ammonia levels in blood had increased approximately 3-fold at 7 days of ammonia ingestion. These changes were associated with a significant decrease in the specific binding of somatostatin (SS) to putative receptors sites in the frontoparietal cortex and hippocampus at 7 and 15 days after starting the high ammonia diet. Scatchard analysis shows that the decrease in SS binding resulted from a decrease in the number of available SS receptors rather than a change in receptor affinity. No changes in the somatostatin-like immunoreactivity content (SSLI) were detected in either brain area at the three study times. These results suggest that hyperammonemia alone can affect the rat brain somatostatinergic system. However, the animal model of hyperammonemia used here is insufficient to produce encephalopathy despite the significant increase in serum ammonia.

Effect of ammonium acetate on the somatostatinergic system in the rat frontoparietal cortex.

Short (90 min)-, mid (5 days)-, and long-term (15 days) ammonium acetate (5 mmol/kg IP) administration decreased the number of specific [125I][Tyr11]somatostatin receptors in synaptosomes from the frontoparietal cortex without changing the affinity constant. Administration of ammonium acetate did no affect the levels of somatostatin-like immunoreactivity in the frontoparietal cortex. The administration of a single dose of N-carbamyl-L-glutamate (1 mmol/kg) plus L-arginine (1 mmol/kg) 1 h before the last administration of ammonium acetate totally blocked the inhibitory effects of the latter on somatostatin receptor number in the frontoparietal cortex synaptosomes. N-Carbamyl-L-glutamate plus L-arginine alone had no observable effect on the somatostatinergic system. The decrease in the number of somatostatin receptors induced by ammonium acetate might reflect decreased target cell sensitivity to somatostatin, a phenomenon that could contribute to the depressed neuronal excitability induced by ammonia in the rat frontoparietal cortex.

Immunohistochemical localization and distribution of VIP/PACAP receptors in human lung.

VIP and PACAP are distributed in nerve fibers throughout the respiratory tract acting as potent bronchodilators and secretory agents. By using RT-PCR and immunoblotting techniques, we have previously shown the expression of common VIP/PACAP (VPAC(1) and VPAC(2)) and specific PACAP (PAC(1)) receptors in human lung. Here we extend our aims to investigate by immunohistochemistry their localization and distribution at this level. A clear immunopositive reaction was obtained in human lung sections by using either anti-VPAC(1) or -VPAC(2) receptor antibodies but not with anti-PAC(1) receptor antibody. However, PAC(1) receptor (and VPAC(1) and VPAC(2) receptors) could be identified in lung membranes by immunoblotting which supports that the PAC(1) receptor is expressed at a low density. Both VPAC(1) and VPAC(2) receptors showed similar immunohistochemical patterns appearing in smooth muscle cells in the wall of blood vessels and in white blood cells (mainly in areas with inflammatory responses). The results agree with previous evidence on the importance of both peptides in the immune system and support their anti-inflammatory and protective roles in lung.

Hyperammonemia induces transient GFAP immunoreactivity changes in goldfish spinal cord (Carassius auratus L.).

In this study we have demonstrated that high ammonia concentration in tank water induces changes in the glial fibrillary acidic protein (GFAP) of ependymal cells and radial astrocytes in the goldfish spinal cord. Hyperammonemia was induced by elevating the ammonia concentration in the tank water to 0.88 mM using ammonium chloride; ammonia in control water was less than 0.1 mM. Immunohistochemical methods were used for GFAP and vimentin, and levels were measured at 4, 8, 16, 30, 60, 90 and 120 days. GFAP quantification was made by means of a digital analysis system. The GFAP immunoreactivity was significantly lower at 30 and 60 days of treatment and at 90 days it had returned to control levels. However, no changes in vimentin immunoreactivity were appreciated in any case. GFAP loss was general and was not selective in any specific spinal cord region. To explain this transient generalized loss of GFAP and its posterior recuperation, a possible relation between glutamine synthetase distribution and GFAP changes is discussed.

Effects of experimentally induced hyperammonemia on glial fibrillary acidic protein (GFAP) in the rhombencephalon of goldfish (Carassius auratus L.).

This experimental study was made to know the effect of hyperammonemia on glial fibrillary acidic protein (GFAP) in the glial cells of posterior rhombencephalon in the goldfish (Carassius auratus L.). Hyperammonemia was induced by elevating the ammonia concentration in the tank water to 0.88 mM with ammonium chloride; the ammonia level in the control tank water was < 0.1 mM. The GFAP levels were measured at 8, 16, 30, 60, 90 and 120 days. GFAP was quantified with a digital analysis system and a transient heterogeneous decrease of GFAP was observed. Hyperammonemia mostly affected GFAP in the astrocyte processes associated with cholinergic pathways. An explanation for the adaptive response to hyperammonemia by fish astrocytes is suggested.

Heat-shock proteins expression in fish central nervous system and its possible relation with water acidosis resistance.

The expression of 70 and 60-kDa heat-shock proteins (HSP70 and HSP60) and glial fibrillary acidic protein (GFAP), determined by immunoblotting and immunohistochemical methods, was studied in fish neural tissue; moreover the possible correlation between the expression of these proteins in neural tissue and fish acidosis resistance was also examined. The HSP GFAP content was analyzed in four different teleostean fish species (gourami, carp, goldfish and trout) under control conditions and in carp under experimental conditions to induce HSPs expression. Under control conditions, HSP70 and HSP60 expression was similar in gourami, carp and goldfish, but gourami had the highest acidosis resistance; trout had the lowest HSP70 and 60 expression and lowest acidosis resistance. The HSP expression pattern was mainly neuronal under control conditions. HSP expression was induced in carp and the effect of this induction on acidosis resistance was studied. Two methods were used for HSP induction in carp: acid shock (2 h at 4.5 pH) and heat shock (2 h at 33 degrees C). A high acidosis resistance, although non-significant, was observed after heat pretreatment. An important HSP expression was detected in glial cells after induction. GFAP expression showed no association with acidosis resistance under either control or experimental conditions.

Modulation of AMPA receptor subunits GluR1 and GluR2/3 in the rat cerebellum in an experimental hepatic encephalopathy model.

The immunohistochemical expression and distribution of the AMPA-selective receptor subunits GluR1 and GluR2/3 were investigated in the rat cerebellum following portocaval anastomosis (PCA) at 1 and 6 months. With respect to controls, GluR1 and GluR2/3 immunoreactivities increased over 1 to 6 months following PCA, although immunolabelling patterns for both antibodies were different at the two analysed times. GluR1 immunoreactivity was expressed by Bergmann glial cells, which showed immunoreactive glial processes crossing the molecular layer at 6 months following PCA. The GluR2/3 subunit was expressed by Purkinje neurons and moderately expressed by neurons of the granule cell layer. Immunoreactivity for GluR2/3 was detectable in cell bodies and dendrites of Purkinje cells in young control cerebella, whereas GluR2/3 immunoreactivity was scarce 1 month post PCA. However, despite a lack of immunoreactivity in the Purkinje somata and main processes of adult control rats, GluR2/3 immunoreactivity was strongly enhanced in Purkinje neurons following long-term PCA. These findings suggest that the localization of the GluR2/3 subunit in Purkinje cells undergoes an alteration and/or reorganization as a consequence of long-term PCA. The combination of enhanced GluR immunoreactivity in long-term PCA, both in Bergmann glial cells and in Purkinje neurons, suggests some degree of neuro-glial interaction, possibly through glutamate receptors, in this type of encephalopathy.

Modulation of glutamate transporters (GLAST, GLT-1 and EAAC1) in the rat cerebellum following portocaval anastomosis.

Glutamate transporters have the important function of removing glutamate released from synapses and keeping extracellular glutamate concentrations below excitotoxic levels. Extracellular glutamate increases in portocaval anastomosis (PCA), so we used a portacaval anastomosis model in rats to analyze the expression of glutamate transporters (GLAST, GLT-1 and EAAC1) in rat cerebellum, 1 and 6 months after PCA, using immunohistochemical methods. In controls, EAAC1 immunoreactivity in Purkinje cells and glial GLAST and GLT-1 immunoreactivities in the molecular layer (ML) increased from young to old rats. One month after PCA, Purkinje cell bodies were not immunostained for neuronal EAAC1 glutamate transporter, whereas glial glutamate transporter expressions (GLAST and GLT-1) were decreased when compared to young controls. In rats with long-term PCA (6 months post-PCA), neuronal and glial glutamate transporter expressions were increased. The expression of the neuronal glutamate transporter EAAC1 was less intense than old controls, whereas glial glutamate transporters (GLAST and GLT-1) increased more than their controls. Since the level of the neuronal glutamate transporter (EAAC1) in long-term PCA did not reach that of the controls, GLAST and GLT-1 glutamate transporters seemed to be required to ensure the glutamate uptake in this type of encephalopathy. EAAC1 immunoreactivity also was expressed by Bergmann glial processes in long-term PCA, but this increase did not suffice to reverse the alterations caused at the early stage. The present findings provide evidence that transitory alteration of glutamate transporter expressions could be a significant factor in the accumulation of excess glutamate in the extracellular space in PCA, which probably makes Purkinje cells more vulnerable to glutamate effect.

The benzodiazepine antagonist CGS 8216 prevents hyperammonemia-induced somatostatin receptor reduction in the brain.

Previous results from our group showed that hyperammonemia decreases the number of somatostatin (SS) receptors and that benzodiazepine receptors might regulate the number of SS receptors in rat brain. These findings together with the supersensitivity of benzodiazepine receptors in the hyperammonemic rat brain suggest that benzodiazepine receptors might mediate the effect of hyperammonemia on SS receptors. To assess this hypothesis we tested whether 2-phenylpyrazolo[3,4-c]-quinolin-3(5H)-one (CGS 8216), a benzodiazepine antagonist, prevented the effect of ammonium acetate on rat brain SS receptors. Administration of ammonium acetate (5 mmol/kg, i.p.) for 7 days did not affect the levels of somatostatin-like immunoreactivity but decreased the number of SS receptors in synaptosomes from the frontoparietal cortex and hippocampus without affecting their apparent affinity. This decrease could be blocked by the concomitant administration of CGS 8216 (10 mg/kg, i.p.). The benzodiazepine antagonist alone had no observable effect on the somatostatinergic system. These results suggested that the effect of hyperammonemia on SS receptors could be mediated, at least in part, through the benzodiazepine receptors.

Postnatal development of glial fibrillary acidic protein immunoreactivity in the hamster arcuate nucleus.

The postnatal development (from 2 days to 1 year) of glial fibrillary acidic protein (GFAP) immunoreactive cells was studied in the arcuate nucleus of male hamsters. In the first postnatal week, GFAP immunoreactivity was observed in radial glial cells whose cell bodies were located in the ependymal layer. Cell processes of GFAP immunoreactive radial glia crossed the arcuate nucleus and reached the pial surface, where they formed a thin and incomplete external limiting membrane. During the second postnatal week, some immunoreactive cell bodies were also located far from the ependymal layer. Some of these cell bodies presented processes that made contact with the ependymal layer whereas others, probably corresponding to maturing astrocytes, did not show ventricular connections. In the third week, only astrocytes showed GFAP immunoreactive perikarya and their immunoreactive processes reached either the blood vessels to form end-feet, or the basal hypothalamic zone to form the glia limitans. In successive weeks, there was an increase of the amount of GFAP-immunoreactive profiles on the glia limitans and surrounding the arcuate nucleus blood vessels. After the 6th postnatal week we observed some GFAP-immunoreactive cells close to arcuate neurons. The number of these cells increased from the 8th postnatal week. From this age on GFAP immunoreactive astrocytic processes compartimentalized the arcuate nucleus defining several rows of aligned neurons. These results indicate that the cytoarchitectonic organization of GFAP immunoreactive elements and their relationship with neurons, blood vessels and pia is not completed until the first 8 weeks of postnatal life in the arcuate nucleus of the hamster.

Fine structural relationships between astrocytes and the node of Ranvier in the amphibian and reptilian spinal cord.

Associations between the axolemma of nodes of Ranvier and perinodal astrocytes in the amphibian and reptilian spinal cord were examined by electron microscopy. Our present results demonstrate the astrocytic processes at the nodes of Ranvier in the central nervous system of lower vertebrates. Such a presence in lower vertebrates is discussed in the framework of current views on neuron-glial interactions and their possible phylogenetic implications.

Glial fibrillary acidic protein and vimentin immunohistochemistry in the posterior rhombencephalon of the Iberian barb (Barbus comiza).

The gustatory centers (vagal lobes and facial lobe) and the tegment of the posterior rhombencephalon of the Iberian barb (Barbus comiza) have been studied using anti-glial fibrillary acidic protein (anti-GFAP) and anti-vimentin immunohistochemical techniques. GFAP immunoreactivity was found in the tegment and in a part of the vagal lobes while vimentin immunoreactivity was located in the tegment. Two immunopositive cell types were found: ependymocytes and radial astrocytes. Since the distribution of GFAP in the barb rhombencephalon corresponds with zones previously described as cholinergic, the GFAP-immunopositive radial astrocytes might be involved in acetylcholine metabolism.

The somatostatin receptor-adenylate cyclase system in rat pancreatic acinar membranes after temporary pancreaticobiliary duct ligation.

The mechanism whereby somatostatin (SS) produces beneficial effects in established pancreatitis induced by pancreaticobiliary duct ligation (PBDL) is still not clear. The aim of the work was to evaluate the possibility of a direct action of SS on pancreatic acinar cells from rats with acute pancreatitis. For this purpose, we studied the SS-receptor-adenylate cyclase system in pancreatic acinar membranes from both, control rats and rats with experimentally induced acute pancreatitis. On the other hand, it has been reported that cholecystokinin (CCK) diminishes the number of SS receptors in pancreatic acinar cells. Proglumide, a CCK receptor antagonist reduces the severity of acute pancreatitis in the rat. Therefore, we have also examined the effect of proglumide on the somatostatinergic system in controls and rats with acute pancreatitis. Fourteen hours after PBDL, the SS receptors, the capacity of the SS analogue SMS 201-995 to inhibit forskolin-stimulated adenylate cyclase activity and PTX-catalyzed [32P] ADP-ribosylation of the alpha1 subunits of Gi proteins could not be detected in pancreatic acinar membranes. One month after reopening the closed pancreaticobiliary duct (PBD), the pancreas showed regeneration of acinar cells, and the above-mentioned parameters were significantly lower than in the control group. Two months after reopening the closed PBD, all these parameters had returned to control values. The administration of proglumide (20 mg/kg i.p.), a cholecystokinin receptor antagonist, accelerated pancreatic regeneration and approached all these parameters to control values one month after reopening the closed PBD. The present study suggests that the beneficial effects of SS on established pancreatitis induced by PBDL may not be due to a direct action of the peptide on pancreatic acinar cells at least at 14 hours after PBDL. In addition, these findings suggest that in established pancreatitis the effect of proglumide on the SS receptor-adenylate cyclase system could be due to its action on pancreatic regeneration.