Diabetes enhances oxidative stress-induced TRPM2 channel activity and its control by N-acetylcysteine in rat dorsal root ganglion and brain

N-acetylcysteine (NAC) is a sulfhydryl donor antioxidant that contributes to the regeneration of glutathione (GSH) and also scavengers via a direct reaction with free oxygen radicals. Recently, we observed a modulatory role of NAC on GSH-depleted dorsal root ganglion (DRG) cells in rats. NAC may have a protective role on oxidative stress and calcium influx through regulation of the TRPM2 channel in diabetic neurons. Therefore, we investigated the effects of NAC on DRG TRPM2 channel currents and brain oxidative stress in streptozotocin (STZ)-induced diabetic rats. Thirty-six rats divided into four groups: control, STZ, NAC and STZ + NAC. Diabetes was induced in the STZ and STZ + NAC groups by intraperitoneal STZ (65 mg/kg) administration. After the induction of diabetes, rats in the NAC and STZ + NAC groups received NAC (150 mg/kg) via gastric gavage. After 2 weeks, DRG neurons and the brain cortex were freshly isolated from rats. In whole-cell patch clamp experiments, TRPM2 currents in the DRG following diabetes induction with STZ were gated by H₂O₂. TRPM2 channel current densities in the DRG and lipid peroxidation levels in the DRG and brain were higher in the STZ groups than in controls; however, brain GSH, GSH peroxidase (GSH-Px), vitamin C and vitamin E concentrations and DRG GSH-Px activity were decreased by diabetes. STZ + H₂O₂-induced TRPM2 gating was totally inhibited by NAC and partially inhibited by N-(p-amylcinnamoyl) anthranilic acid (ACA) and 2-aminoethyl diphenylborinate (2-APB). GSH-Px activity and lipid peroxidation levels were also attenuated by NAC treatment. In conclusion, we observed a modulatory role of NAC on oxidative stress and Ca²⁺ entry through the TRPM2 channel in the diabetic DRG and brain. Since excessive oxidative stress and overload Ca²⁺ entry are common features of neuropathic pain, our findings are relevant to the etiology and treatment of pain neuropathology in DRG neurons.     

Gastrin reverses established cholangiocyte proliferation and enhanced secretin‐stimulated ductal secretion of BDL rats by activation of apoptosis through increased expression of Ca2+‐dependent PKC isoforms

Abstract: We posed these questions: (i) Does administration of gastrin to 1‐week bile duct ligation (BDL) rats inhibits established cholangiocyte proliferation and ductal secretion? (ii) Is gastrin inhibition of cholangiocyte proliferation and secretion of BDL rats associated with enhanced apoptosis? (iii) Are gastrin's effects on cholangiocyte function associated with increased expression of protein kinase C (PKC) isoforms; and (iv) Is gastrin stimulation of cholangiocyte apoptosis regulated by the Ca²⁺‐dependent PKC pathway? Methods: Seven days after BDL, rats were treated with gastrin by minipumps for 14 days. Cholangiocyte proliferation was assessed by measurement of the number of PCNA and CK‐19 positive cholangiocytes in sections, and PCNA expression in cholangiocytes. Ductal secretion was determined by measurement of secretin‐induced cAMP levels and choleresis. Apoptosis was evaluated by TUNEL analysis in sections and annexin‐V staining in cholangiocytes. The expression of PKC isoforms was determined by immunoblots. Results: Gastrin inhibits established cholangiocyte proliferation and enhanced secretin‐stimulated ductal secretion of BDL rats. Gastrin's effects on cholangiocyte function were associated with enhanced apoptosis and increased expression of PKC alpha, and beta I and II. Gastrin increases in cholangiocyte apoptosis were blocked by BAPTA/AM and H7. Summary/conclusion: Gastrin inhibits cholangiocyte proliferation and secretin‐induced ductal secretion in BDL rats by increasing apoptosis through a PKC‐mediated mechanism.