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

Abstract

SUMMARY1. Desensitization of the Ca2+ release response evoked by acetylcholine in acinar cells from rat lacrimal glands was studied using the Ca2+-sensitive dye Fura-2. The evolution of the amplitude and half-maximal rise time (t1) 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 ti with trial number. Both amplitude and ti recovered their control values after 20 min of washing.3. Staurosporine (02-1 ,lM), an inhibitor of protein kinase C, was found to decrease the slopes of the variation of amplitude and ti 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 /LM), 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 ICM) 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.

Role of protein kinase C in cholinergic stimulation of lacrimal gland protein secretion

“…Moreover, PdBu treatment increased phenylephrine-induced peroxidase secretion [24]. These results strongly suggest that PKC activity is effectively downregulated by phorbol ester treatment and are in agreement with Tan and Marty [25] who reported that a 224 h treatment of lacrimal gland acinar cells with PMA down-regulated PKC.…”

Neural regulation of lacrimal gland secretory processes: Relevance in dry eye diseases

“…Activation of PKC can alter the Ca 2+ influx pathway as inhibition of PKC with the non-selective PKC inhibitor staurosporine decreased the plateau Ca 2+ in response to cholinergic agonists and preactivation of cPKC and nPKC isoforms with phorbol esters decreased both the peak and plateau Ca 2+ response to cholinergic agonists (Zoukhri, Hodges et al 2000). This is in agreement with the work of Tan and Marty (Tan and Marty 1991), Berrie and Elliot (Berrie and Elliott 1994), and Gromada et al (Gromada, Jorgensen et al 1995) that PKC isoforms decrease the production of InsP 3 , as well as the Ca 2+ influx pathway. Using the myristoylated pseudosubstrate inhibitors, Zoukhri et al found that nPKCδ and nPKCε, but not cPKCα, inhibited Ca 2+ influx (Zoukhri, Hodges et al 2000).…”

Intracellular alkalinization mobilizes calcium from agonist‐sensitive pools in rat lacrimal acinar cells.

“…Since evoked a larger Cr release than 20 mm cell was stimulated twice more with ACh in ssion. This repeated stimulation produces on of the ACh response, as previously reported and others (Tan & Marty, 1991 initial increase in [Ca2+]i comparable to that evoked by 20 mm NH4Cl in the same cell. However, it was clear that, in contrast to NH4Cl, ACh also produced a sustained elevation of [Ca2+]i for as long as it was applied.…”