Effect of phospholipase C from Bacillus cereus on the release of membrane-bound choline-O-acetyltransferase from rat hippocampal tissue.
Some of the enzyme choline-O-acetyltransferase (ChAT) associated with central cholinergic nerve terminals appears to be non-ionically associated with membranes. In the present study, we tested the possibility that some membrane-bound ChAT might be anchored to membranes by a phosphatidylinositol linkage by incubating rat hippocampal tissue with phospholipase C (PLC) from Bacillus cereus. The PLC selectively augmented the release of ChAT; also, the glycosylphosphatidylinositol-PLC inhibitor, zinc, blocked this increase in release. When control and PLC-treated hippocampal tissues were subjected to Triton X-114 phase separation, a procedure that separates amphiphilic from hydrophilic proteins, the detergent-soluble, membrane-bound fraction of tissue ChAT appeared to be the source of the ChAT released by PLC into the incubation medium. Zinc also blocked the temperature-dependent release of ChAT, but not lactic dehydrogenase, from hippocampal tissue. Extracellular membrane-bound ChAT appeared to be the source of the ChAT released by a low exogenous concentration of PLC, as well as that released by a temperature-dependent process during tissue incubation. Phosphatidylinositol-specific PLC from Bacillus thuringiensis released ChAT, but not lactic dehydrogenase, from a crude synaptosomal fraction prepared from rat hippocampal tissue. These results suggest that some of the membrane-bound ChAT in rat hippocampal tissue may be extracellular and anchored to the membrane by phosphatidylinositol, and also that an endogenous factor in hippocampal tissue may function to remove this extracellular ChAT from the membrane.
Carroll
P T
PT
Department of Pharmacology, Texas Tech University Health Sciences Center, Lubbock 79430.
Smith
L K
LK
D000818:Animals; D001409:Bacillus cereus / Q000201:enzymology*; D002794:Choline / Q000494:pharmacology; D002795:Choline O-Acetyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003902:Detergents; D005110:Extracellular Space / Q000378:metabolism; D006624:Hippocampus / Q000201:enzymology*; D006652:Histological Techniques; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008297:Male; D008566:Membranes / Q000201:enzymology; D009994:Osmolar Concentration; D010716:Phosphatidylinositols / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D012995:Solubility; D013696:Temperature; D010738:Type C Phospholipases / Q000494:pharmacology*; D015032:Zinc / Q000494:pharmacology
D003902:Detergents; D010716:Phosphatidylinositols; D007770:L-Lactate Dehydrogenase; D002795:Choline O-Acetyltransferase; D010738:Type C Phospholipases; D015032:Zinc; D002794:Choline