Caffeine-induced calcium release from internal stores in cultured rat sensory neurons.
Free intracellular calcium concentration ([Ca2+]in) was recorded at 22 degrees C by means of Indo-1 or Fura-2 single-cell microfluorometry in cultured dorsal root ganglion neurons obtained from neonatal rats. The resting [Ca2+]in in dorsal root ganglion neurons was 73 +/- 21 nM (mean +/- S.D., n = 94). Fast application of 20 mM caffeine evoked [Ca2+]in transient which reached a peak of 269 +/- 64 nM within 5.9 +/- 1.1 s. After reaching the peak the [Ca2+]in level started to decline in the presence of caffeine and for 87.2 +/- 10.6 s cytoplasmic calcium returned to an initial resting value. In 40% of neurons tested [Ca2+]in decreased to subresting levels following the washout of caffeine (the so-called post-caffeine undershoot). On average, the undershoot level was 19 +/- 2.5 nM below the resting [Ca2+]in value. Prolonged exposure of caffeine depleted the caffeine-sensitive stores of releasable Ca2+; the degree of this depletion depended on caffeine concentration. The depletion of the caffeine-sensitive internal stores to some extent was linked to calcium extrusion via La(3+)-sensitive plasmalemmal Ca(2+)-ATPases. The stores could be partially refilled by the uptake of cytoplasmic Ca2+, but the complete recovery of releasable Ca2+ content of the caffeine-sensitive pools required the additional calcium entry via voltage-operated calcium channels. Caffeine-evoked [Ca2+]in transients were effectively blocked by 10 microM ryanodine, 5 mM procaine, 10 microM dantrolene or 0.5 mM Ba2+, thus sharing the basic properties of the Ca(2+)-induced-Ca2+ release from endoplasmic reticulum. Pharmacological manipulation with caffeine-sensitive stores interfered with the depolarization-induced [Ca2+]in transients. In the presence of low caffeine concentration (0.5-1 mM) in the extracellular solution the rate of rise of the depolarization-triggered [Ca2+]in transients significantly increased (by a factor 2.15 +/- 0.29) suggesting the occurrence of Ca(2+)-induced Ca2+ release. When the caffeine-sensitive stores were emptied by prolonged application of caffeine, the amplitude and the rate of rise of the depolarization-induced [Ca2+]in transients were decreased. These facts suggest the involvement of internal caffeine-sensitive calcium stores in the generation of calcium signal in sensory neurons.
Usachev
Y
Y
Bogomoletz Institute of Physiology, Kiev, Ukraine.
Shmigol
A
A
Pronchuk
N
N
Kostyuk
P
P
Verkhratsky
A
A
D000818:Animals; D000831:Animals, Newborn; D001464:Barium / Q000494:pharmacology; D002110:Caffeine / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002478:Cells, Cultured; D005727:Ganglia, Spinal / Q000166:cytology / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism*; D009475:Neurons, Afferent / Q000378:metabolism*; D009994:Osmolar Concentration; D011343:Procaine / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D012433:Ryanodine / Q000494:pharmacology
D002121:Calcium Channel Blockers; D012433:Ryanodine; D001464:Barium; D002110:Caffeine; D011343:Procaine; D002118:Calcium