Role of intracellular calcium in fast and slow desensitization of P₂-receptors in PC12 cells

Abstract

1. Combined whole-cell patch clamp recording and confocal laser scanning microscopy of [Ca²⁺]_i transients were performed on single PC12 cells to study any correlation between membrane currents induced by ATP and elevation in [Ca²⁺]_i. ATP was applied by pressure from micropipettes near the recorded PC12 cells continuously superfused at a fast rate.

2. Brief (20 ms) pulses of ATP elicited monophasic inward currents and [Ca²⁺]_i increases. Long applications (2 s) of ATP (5 mM) evoked peak currents which rapidly faded during the pulse and were followed by a large rebound current, interpreted as due to rapid desensitization and recovery of P₂- receptors. The associated [Ca²⁺]_i increase grew monotonically to a peak reached only after the occurrence of the current rebound, indicating that it is unlikely this cation has a role in fast desensitization.

3. Both membrane currents and [Ca²⁺]_i transients were linearly dependent on holding membrane potential, suggesting that Ca²⁺ influx is the predominant cause of [Ca²⁺]_i elevation. This view was supported by experiments carried out in Ca²⁺-free solution.

4. Brief pulses of ATP applied after a desensitizing pulse (2 s) of the same elicited smaller inward currents and [Ca²⁺]_i rises indicating a role for [Ca²⁺]_i in controlling slow desensitization of P₂-receptors.

5. This notion was confirmed in experiments with various [Ca²⁺]_i chelators which differentially affected slow desensitization in relation to their buffering capacity, while sparing fast receptor desensitization.

6. These results suggest a role for [Ca²⁺]_i in slow rather than fast desensitization of P₂-receptors, thus proposing this divalent cation as an intracellular factor able to provide an efficient and reversible control over receptor activity induced by ATP.

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