Evidence for two modes of Ca2+ entry following muscarinic stimulation of a human salivary epithelial cell line.
We have investigated muscarinic receptor-operated Ca2+ mobilization in a salivary epithelial cell line. HSG-PA, using an experimental approach which allows independent evaluation of intracellular Ca2+ release and extracellular Ca2+ entry. The carbachol (Cch) dose response of intracellular Ca2+ release indicates the involvement of a single, relatively low-affinity, muscarinic receptor site (K0.5 approximately equal to 10 or 30 microM, depending on the method for [Ca2+]i determination). However, similar data for Ca2+ entry indicate the involvement of two Cch sites, one consistent with that associated with Ca2+ release and a second higher affinity site with K0.5 less than or equal to 2.5 microM. In addition, the Ca2+ entry response observed at lower concentrations of Cch (2.5 microM) was completely inhibited by membrane depolarization induced with high K+ (greater than 55 mM) or gramicidin D (1 microM), while membrane depolarization had little or no effect on Ca2+ entry induced by 100 microM Cch. Another muscarinic agonist, oxotremorine-M (100 microM; Oxo-M), like Cch, also induced an increase in the [Ca2+]i of HSG-PA cells (from 72 +/- 2 to 104 +/- 5 nM). This response was profoundly blocked (approximately 75%) by the inorganic Ca2+ channel blocker La3+ (25-50 microM) suggesting that Oxo-M primarily mobilizes Ca2+ in these cells by increasing Ca2+ entry. Organic Ca2+ channel blockers (verapamil or diltiazem at 10 microM, nifedipine at 1 microM), had no effect on this response. The Oxo-M induced Ca2+ mobilization response, like that observed at lower doses of Cch, was markedly inhibited (approximately 70-90%) by membrane depolarization (high K+ or gramicidin D).(ABSTRACT TRUNCATED AT 250 WORDS)
He
X J
XJ
Clinical Investigations and Patient Care Branch, National Institute of Dental Research, National Institutes of Health, Bethesda, Maryland 20892.
Wu
X Z
XZ
Turner
R J
RJ
Baum
B J
BJ
D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology*; D002460:Cell Line; D004848:Epithelium / Q000378:metabolism; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008564:Membrane Potentials; D010095:Oxotremorine / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000378:metabolism*; D012469:Salivary Glands / Q000166:cytology / Q000378:metabolism*
D002121:Calcium Channel Blockers; D011976:Receptors, Muscarinic; D010095:Oxotremorine; D015544:Inositol 1,4,5-Trisphosphate; D002217:Carbachol; D002118:Calcium