Abstract
The free intracellular Ca2+ concentration ([Ca2+](i)) is governed by the balance between the activation of Ca2+ channels and buffering and efflux processes. We tested the hypothesis that Ca2+ efflux pathways are susceptible to modulation. The whole-cell patch-clamp technique was used in combination with Indo-1-based microfluorometry to record Ca2+ current and [Ca2+](i) simultaneously from single rat dorsal root ganglion (DRG) neurons grown in culture. Depolarizing test pulses (-80 to 0 mV, 100-300 msec) elicited [Ca2+](i) transients that recovered to basal levels by a process best-fit with a single exponential (τ = 5.1 ± 0.4 sec; n = 14) and were independent of Ca2+ load (40-500 pC) over this range of test pulses. [Ca2+](i) transients recorded in whole-cell configuration were similar to those elicited by a brief train of action potentials in unclamped neurons. Inhibition of Ca2+ sequestration into intracellular stores with thapsigargin had no effect on the kinetics of recovery. Inhibition of plasma membrane Ca2+ ATPase (PMCA) function by including a peptide inhibitor (C28R2) in the patch pipette significantly slowed recovery to basal [Ca2+](i) (τ = 9.9 ± 0.8 sec; n = 4). Preincubation with calmidazolium, a calmodulin antagonist, produced modest slowing of Ca2+ efflux. Phorbol dibutyrate, an activator of protein kinase C (PKC), accelerated Ca2+ efflux only when the PMCA had been inhibited by C28R2. We conclude that in DRG neurons PMCAs are responsible for lowering [Ca2+](i) after small Ca2+ loads and that PMCA-mediated Ca2+ efflux is modulated by calmodulin- and PKC-signaling pathways.
Original language | English (US) |
---|---|
Pages (from-to) | 1008-1015 |
Number of pages | 8 |
Journal | Journal of Neuroscience |
Volume | 16 |
Issue number | 3 |
DOIs | |
State | Published - Feb 1 1996 |
Keywords
- Ca ATPase
- Indo- 1
- calmodulin
- dorsal root ganglion
- intracellular calcium
- patch clamp
- protein kinase C