Protein kinase C‐mediated desensitization of the muscarinic response in rat lacrimal gland cells.

1. Desensitization of the Ca2+ release response evoked by acetylcholine in acinar cells from rat lacrimal glands was studied using the Ca(2+)‐sensitive dye Fura‐2. The evolution of the amplitude and half‐maximal rise time (t1/2) of the Ca2+ response was followed as a function of trial number in a series of stimulations. 2. Under control conditions repetitive applications of acetylcholine (15 s long applications every minute) led to a linear decrease of the amplitude and to a linear increase of t1/2 with trial number. Both amplitude and t1/2 recovered their control values after 20 min of washing. 3. Staurosporine (0.2‐1 microM), an inhibitor of protein kinase C, was found to decrease the slopes of the variation of amplitude and t1/2 with trial number. 4. Prolonged treatment with 12‐O‐tetradecanoyl phorbol 13‐acetate (TPA), an activator of protein kinase C (100‐250 nM for 2‐4 h), also led to a markedly decreased desensitization, presumably as a result of down‐regulation of protein kinase C. On the other hand moderate pre‐treatments with TPA (16‐32 nM for 10 min) strongly inhibited the response, most probably as a result of protein kinase C activation. 5. Application of oleoylacetylglycerol (50 microM), a weaker activator of protein kinase C, inhibited the response and enhanced desensitization. These effects were, however, not obtained after down‐regulation of protein kinase C with strong exposure to TPA. 6. We conclude that protein kinase C activation following the ACh‐induced Ca2+ rise and the concomitant diacylglycerol production mediates desensitization of the response. 7. Arachidonic acid (100 microM) inhibited the ACh‐induced response and enhanced desensitization. However, this effect did not appear to be mediated by protein kinase C since it was also obtained with docosahexaenoic acid, an analogue of arachidonic acid which does not activate protein kinase C.