Depolarization- and transmitter-induced changes in intracellular Ca2+ of rat cerebellar granule cells in explant cultures.
Digital imaging of the Ca indicator fura-2 has been used to study the responses of developing granule cells in culture to depolarization and transmitter action. Unstimulated cells bathed in Krebs saline exhibited cytoplasmic Ca ion concentrations, [Ca2+], that were generally in the 30-60 nM range. Exposure of cells to high-potassium (25 mM) saline depolarized the membrane potential and produced an immediate rise in [Ca2+] that recovered within 2-3 min in normal saline. The response grew progressively larger over the first 20 d in culture. Transient increases in [Ca2+] to levels greater than 1 microM were observed after 12-14 d in vitro, at which time the cells displayed intense electrical activity when exposed to high K. At this stage, the increases were attenuated by blocking action potential activity with TTX. In TTX-treated or immature cells, in which the transient phase of the Ca change was relatively small, a second exposure to high K typically produced a much larger Ca response that the initial exposure. The duration of this facilitation of the response persisted for periods longer than 5 min. Application of the neurotransmitter GABA induced a transient increase in membrane conductance, with a reversal potential near resting potential (approx. -60 mV), and caused an intracellular Ca2+ increase that outlasted the exposure to GABA by several minutes. Glutamate, or kainate, induced an increase in membrane conductance but with a reversal potential more positive than spike threshold. These agents also elevated intracellular Ca2+, but unlike the case with GABA, this Ca response reversed rapidly upon removal of the transmitter. The facilitatory effect of repeated exposures to high-K saline, as well as the persistent Ca elevation following a brief GABA application, suggests that granule cells possess the capability of displaying activity-dependent changes in Ca levels in culture.
D000818:Animals; D001572:Benzofurans; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000378:metabolism*; D004553:Electric Conductivity; D005456:Fluorescent Dyes; D016257:Fura-2; D005971:Glutamates / Q000502:physiology*; D018698:Glutamic Acid; D007091:Image Processing, Computer-Assisted; D007608:Kainic Acid / Q000494:pharmacology; D008564:Membrane Potentials; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D013779:Tetrodotoxin / Q000494:pharmacology; D005680:gamma-Aminobutyric Acid / Q000502:physiology*
D001572:Benzofurans; D005456:Fluorescent Dyes; D005971:Glutamates; D018698:Glutamic Acid; D013779:Tetrodotoxin; D005680:gamma-Aminobutyric Acid; D011188:Potassium; D007608:Kainic Acid; D002118:Calcium; D016257:Fura-2