SUMMARY1. Two clones of rat phaeochromocytoma PC12 cells have been used to study the expression of Ca2+ channels and their possible involvement in neuronal differentiation. One clone differentiated morphologically when exposed to nerve growth factor (NGF) for 4 days (PC12 cells), while the other clone was insensitive to NGF, but differentiated morphologically in the presence of ouabain (041 mM

In the central nervous system (CNS), the Na(+)-Ca(2+) exchanger plays a fundamental role in controlling the changes in the intracellular concentrations of Na(+) and Ca(2+) ions. These cations are known to regulate neurotransmitter release, cell migration and differentiation, gene expression, and neurodegenerative processes. In the present study, nonradioactive in situ hybridization and light immunohistochemistry were carried out to map the regional and cellular distribution for both transcripts and proteins encoded by the three known Na(+)-Ca(2+) exchanger genes NCX1, NCX2, and NCX3. NCX1 transcripts were particularly expressed in layers III-V of the motor cortex, in the thalamus, in CA3 and the dentate gyrus of the hippocampus, in several hypothalamic nuclei, and in the cerebellum. NCX2 transcripts were strongly expressed in all hippocampal subregions, in the striatum, and in the paraventricular thalamic nucleus. NCX3 mRNAs were mainly detected in the hippocampus, in the thalamus, in the amygdala, and in the cerebellum. Immunohistochemical analysis revealed that NCX1 protein was mainly expressed in the supragranular layers of the cerebral cortex, in the hippocampus, in the hypothalamus, in the substantia nigra and ventral tegmental area, and in the granular layer of the cerebellum. The NCX2 protein was predominantly expressed in the hippocampus, in the striatum, in the thalamus, and in the hypothalamus. The NCX3 protein was particularly found in the CA3 subregion, and in the oriens, radiatum, and lacunoso-moleculare layers of the hippocampus, in the ventral striatum, and in the cerebellar molecular layer. Collectively, these results suggest that the different Na(+)-Ca(2+) exchanger isoforms appear to be selectively expressed in several CNS regions where they might underlie different functional roles.

Immunocytochemical evidence for localized distribution of the Na+/Ca2+ exchange protein in nerve terminals of cultured hippocampal cells is presented together with results on the functional relevance of the exchanger in the control of [Ca2+]i and of synaptic vesicle recycling. The monoclonal antibody R3F1, directed against an epitope on the intracellular loop of the protein, revealed higher densities of expression in synaptic regions than in other parts of the neurons. Removal of extracellular Na+ produced enhanced and prolonged elevation of [Ca2+]i in nerve terminals during and after electrical stimulation of the cells. Correspondingly, initial rates of exocytosis, measured by fluorescence changes of FM 1-43 during stimulation, were faster in LiCl-containing solution than in NaCl-containing solution. By contrast, endocytosis at 20 s was the same in both solutions.

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SUMMARY1. 45Ca efflux and resting tension were measured in isolated guinea-pig auricles under conditions known to change the intracellular free Ca ion concentration.2. In the presence of [Na]o, caffeine (2 mM) increases 45Ca efflux, but does not produce a contracture, while in the absence of [Na]0 and [Ca]o caffeine causes a contracture without increasing 45Ca efflux. Adrenaline (10-5 -10-4 M) with or without theophylline (0.5-1 0 mM) has no effect on either 45Ca efflux or resting tension.3. In the presence of caffeine the rate of net efflux of Ca depends on [Na]2. Caffeine contractures of muscles in Na-free solution relax upon the addition of [Na]o. Relaxation is correlated with the increase in net efflux of Ca. 4. Cyanide (2 mM) produces a variable increase in 45Ca efflux without a concomitant contracture in Na-containing solutions, but in Na,Ca-free solutions a large contracture occurs without significant increase in 45Ca efflux.5. A large increase in 45Ca efflux and a contracture were observed with the 'Ca-ionophore' X 537A.6. Changes in membrane potential (K-depolarization) in hypertonic solutions have no significant effect on Na-dependent 45Ca efflux, which is in agreement with an electroneutral 2: 1 Na-Ca exchange.7. Cyanide and X 537A both cause a considerable release of Ca ions from isolated guinea-pig heart mitochondria, while caffeine has no effect.