Carbachol-induced phosphatidylcholine hydrolysis and choline efflux in rat submandibular gland involves phospholipase D activation and is modulated by protein kinase C and calcium.
The role of calcium and protein kinase C activation in carbachol-induced choline efflux from submandibular glands was investigated. The participation of phospholipase D in this signal transduction pathway was demonstrated by the formation of [14C]phosphatidylethanol in [14C]lysophosphatidylcholine-labelled submandibular gland cells treated with carbachol or noradrenaline in the presence of ethanol. Chelation of the intracellular calcium with BAPTA/AM reduced the carbachol stimulated outflow of [3H]choline. The calcium ionophore A23187 in a high concentration (10 microM) increased the basal [3H]choline outflow, but decreased the carbachol-induced outflow. Removal of the extracellular calcium enhanced the carbachol-stimulated outflow, which returned to control when calcium was re-added to the medium. Activation of protein kinase C by phorbol-12,13-dibutyrate (100 nM) or 1-oleyl-2-acetyl-sn-glycerol (20 microM) was without effect per se, but enhanced the carbachol-mediated outflow of [3H]choline. Phorbol-12,13-dibutyrate in combination with 1 microM A23187 induced a small efflux of [3H]choline. A 2 h treatment with phorbol-12,13-dibutyrate (1 microM), causing down-regulation of protein kinase C, significantly decreased the carbachol-stimulated [3H]choline outflow. In conclusion, elevation of intracellular calcium levels and protein kinase C activation are of importance for the carbachol-stimulated outflow of [3H]choline. Inflow of calcium, if anything, reduces the carbachol-stimulated outflow of [3H]choline. Since phosphatidylcholine breakdown generates diacylglycerol and this could lead to activation of protein kinase C, activation of this signal transduction pathway may be important for the protein content of the saliva and for the known trophic effects of parasympathetic innervation.
Dunér-Engström
M
M
Department of Physiology and Pharmacology, Karolinska Institute, Sweden.
Fredholm
B B
BB
D000818:Animals; D002118:Calcium / Q000502:physiology; D002217:Carbachol / Q000494:pharmacology*; D002794:Choline / Q000378:metabolism*; D004789:Enzyme Activation; D006868:Hydrolysis / Q000187:drug effects; D008297:Male; D010713:Phosphatidylcholines / Q000378:metabolism*; D010739:Phospholipase D / Q000378:metabolism; D011493:Protein Kinase C / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013363:Submandibular Gland / Q000378:metabolism*
D010713:Phosphatidylcholines; D002217:Carbachol; D011493:Protein Kinase C; D010739:Phospholipase D; D002794:Choline; D002118:Calcium