CYTOPLASMIC CA2+ CONCENTRATIONS IN INTACT MERKEL CELLS OF AN ISOLATED, FUNCTIONING RAT SINUS HAIR PREPARATION

An isolated, functioning sinus hair preparation was developed to inves tigate cytoplasmic Ca2+ concentrations in intact Merkel cells using mi crofluorimetric techniques. Intracellular Ca2+ levels were monitored b y means of photon counters in small groups of Merkel cells loaded with the calcium fluorescent indicators fura-2 or fluo-3. Mechanical stimu lation of Merkel cells with fine glass rods resulted in small transien t increases in intracellular Ca2+ levels (by about 20%) in the group o f Merkel cells around the stimulating probe. A rise in Ca2+ is presume d to be essential for the postulated synaptic transmission to the affe rent nerve terminal. Depolarization with a high concentration of potas sium chloride (100 mM) caused increases in intracellular Ca2+ concentr ations in Merkel cells (by about 70%) only in the presence of extracel lular Ca2+, indicating an influx of Ca2+ through voltage-gated channel s. The Ca2+ response was abolished neither by (+)-BayK8644 nor omega-c onotoxin, suggesting that the Ca2+ channels are different from the cla ssical L- or N-type channels. Extracellular application of ATP (10 mu M to 5 mM) caused dose-dependent increases in intracellular Ca2+ level s in Merkel cells of up to sevenfold from the basal level of about 100 nM. Similar responses to ATP were also measured during superfusion wi th Ca2+-free medium, suggesting intracellular stores as the main Ca2source. Pre-incubation of Merkel cells with the purinoceptor antagonis t suramin (100 mu M) for 30 min reduced the Ca2+ responses to ATP by a bout 50% compared with control conditions. In conclusion, the results have demonstrated that a rise in intracellular Ca2+ in Merkel cells ca n be evoked by mechanical stimulation, membrane depolarization and che mical stimulation by ATP. These observations strongly suggest a possib le contribution of Ca2+ to the normal responsiveness of Merkel cell me chanoreceptors, in turn supporting the hypothesis that Merkel cells ar e involved in the mechano-electric transduction process in sinus hair type I mechanoreceptors.