Regulation of basal intracellular calcium concentration by the sarcoplasmic reticulum in myocytes from the rat gastric antrum

摘要

class="enumerated" style="list-style-type:decimal">The intracellular calcium concentration ([Ca²⁺]i) was monitored in fura-2-loaded myocytes isolated from the rat gastric antrum and voltage clamped at −60 1r1rqmV1qusing the perforated patch clamp technique. The rate of quench of fura-2 fluorescence by Mn²⁺ was used as a measure of capacitative Ca²⁺ entry.Cyclopiazonic acid (5 μM) did not affect the holding current but produced a sustained elevation in steady-state [Ca²⁺]i that was dependent on the presence of external calcium. Cyclopiazonic acid increased Mn²⁺ influx with physiological external [Ca²⁺], but not in Ca²⁺-free conditions. Cyclopiazonic acid increased the rate of [Ca²⁺]i rise following a rapid switch from Ca²⁺-free to physiological [Ca²⁺] solution.Sustained application of carbachol (10 μM) produced an elevation in steady-state [Ca²⁺]i that was associated with an increased rate of Mn²⁺ influx. Application of cyclopiazonic acid in the presence of carbachol further elevated steady-state [Ca²⁺]i without changing Mn²⁺ influx.Ryanodine (10 μM) elevated steady-state [Ca²⁺]i either on its own or following a brief application of caffeine (10 mm). Cyclopiazonic acid had no further effect when added to cells pre-treated with ryanodine. Neither caffeine nor ryanodine increased the rate of Mn²⁺ influx. When brief applications of ionomycin (25 μm) in Ca²⁺-free solution were used to release stored Ca²⁺, ryanodine reduced the amplitude of the resulting [Ca²⁺]i transients by approximately 30 %, indicating that intracellular stores were partially depleted.These findings suggest that continual uptake of Ca²⁺ by the sarcoplasmic reticulum Ca²⁺-ATPase into a ryanodine-sensitive store limits the bulk cytoplasmic [Ca²⁺]i under resting conditions. This pathway can be short circuited by 10 μm ryanodine, presumably by opening Ca²⁺ channels in the sarcoplasmic reticulum. Depletion of stores with cyclopiazonic acid or carbachol also activates capacitative Ca²⁺ entry.