Inositol 1,4,5-trisphosphate receptor isoform expression in mouse pancreatic islets: effects of carbachol.
The inositol 1,4,5-trisphosphate receptors (IP3Rs) are ligand-gated Ca2+ channels that regulate intracellular Ca2+ mobilization. Among the IP3R mRNA isoforms I, II, and III, IP3R-I mRNA was expressed in mouse islets and the beta-cell line betaTC3, and was quantitatively the most abundant isoform as determined by reverse transcriptase-polymerase chain reaction. IP3R-II and -III mRNAs were expressed at similar levels in mouse islets, but neither isoform was detected in betaTC3 cells. Culture of mouse islets for 30 min and 2 hr at 20 mM glucose, or for 7 days at 11 mM glucose did not affect IP3R-I mRNA expression compared with islets cultured in 5.5 mM glucose. Culture of islets or betaTC3 cells with carbachol (0.5 mM) reduced IP3R-I mRNA expression levels below control. Mouse islet alpha- and beta-cells expressed IP3R-I and -III proteins, but IP3R-II protein was not detected by immunoblot or double-label immunohistochemistry. Culture of islets for up to 6 hr with carbachol reduced IP3R-I and -III protein expression in a time-dependent manner with a half-maximal effect on type I at 1 hr. Glucose (20 mM) stimulation for 2 hr did not affect IP3R-1 levels. The carbachol-induced decrease in IP3R-I and -III protein expression was reversed by carbobenzoxyl-leucinyl-leucinyl-leucinyl-H (MG-132), a proteasome inhibitor. Thus, glucose failed to regulate mouse islet IP3R mRNA expression, whereas carbachol stimulation down-regulated IP3R mRNA and protein. A proteasomal protein degradative pathway appeared to mediate the muscarinic receptor-induced effects on IP3R-I and -III.
Lee
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Department of Pharmacology and Toxicology, 102 Farber Hall, State University of New York at Buffalo, School of Medicine and Biomedical Sciences, 14214, Buffalo, NY, USA.
Laychock
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SG
D000704:Analysis of Variance; D000818:Animals; D015220:Calcium Channels / Q000096:biosynthesis* / Q000235:genetics; D002217:Carbachol / Q000494:pharmacology*; D018679:Cholinergic Agonists / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D007150:Immunohistochemistry; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007340:Insulinoma / Q000473:pathology; D007515:Islets of Langerhans / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D020033:Protein Isoforms / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000187:drug effects; D018160:Receptors, Cytoplasmic and Nuclear / Q000096:biosynthesis* / Q000235:genetics; D014407:Tumor Cells, Cultured
D015220:Calcium Channels; D018679:Cholinergic Agonists; D053496:Inositol 1,4,5-Trisphosphate Receptors; D020033:Protein Isoforms; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D002217:Carbachol