Increases in postsynaptic [Ca2+]i can result from Ca2+ entry through ligand-gated channels or voltage-gated Ca2+ channels, or through release from intracellular stores. Most attention has focused on entry through the N-methyl-D-aspartate (NMDA) receptor in causing [Ca2+]i increases since this pathway requires both presynaptic stimulation and postsynaptic depolarization, making it a central component in models of synaptic plasticity. Here, we report that repetitive synaptic activation of metabotropic glutamate receptors (mGluRs), paired with backpropagating action potentials, causes large, wave-like increases in [Ca2+]i predominantly in restricted regions of the proximal apical dendrites and soma of hippocampal CA1 pyramidal neurons. [Ca2+]i changes of several micromolars can be reached by regenerative release caused by the synergistic effect of mGluR-generated inositol 1,4,5-trisphosphate (IP3) and spike-evoked Ca2+ entry acting on the IP3 receptor.

We examined the properties of [Ca(2+)](i) changes that were evoked by backpropagating action potentials in pyramidal neurons in hippocampal slices from the rat. In the presence of the metabotropic glutamate receptor (mGluR) agonists t-ACPD, DHPG, or CHPG, spikes caused Ca(2+) waves that initiated in the proximal apical dendrites and spread over this region and in the soma. Consistent with previously described synaptic responses (Nakamura et al., 1999a), pharmacological experiments established that the waves were attributable to Ca(2+) release from internal stores mediated by the synergistic effect of receptor-mobilized inositol 1,4, 5-trisphosphate (IP(3)) and spike-evoked Ca(2+). The amplitude of the changes reached several micromoles per liter when detected with the low-affinity indicators fura-6F, fura-2-FF, or furaptra. Repetitive brief spike trains at 30-60 sec intervals generated increases of constant amplitude. However, trains at intervals of 10-20 sec evoked smaller increases, suggesting that the stores take 20-30 sec to refill. Release evoked by mGluR agonists was blocked by MCPG, AIDA, 4-CPG, MPEP, and LY367385, a profile consistent with the primacy of group I receptors. At threshold agonist concentrations the release was evoked only in the dendrites; threshold antagonist concentrations were effective only in the soma. Carbachol and 5-HT evoked release with the same spatial distribution as t-ACPD, suggesting that the distribution of neurotransmitter receptors was not responsible for the restricted range of regenerative release. Intracellular BAPTA and EGTA were approximately equally effective in blocking release. Extracellular Cd(2+) blocked release, but no single selective Ca(2+) channel blocker prevented release. These results suggest that IP(3) receptors are not associated closely with specific Ca(2+) channels and are not close to each other.

Calcium-induced calcium release (CICR) is a mechanism by which local elevations of intracellular calcium (Ca2+) are amplified by Ca2+ release from ryanodine-sensitive Ca2+ stores. CICR is known to be coupled to Ca2+ entry in skeletal muscle, cardiac muscle, and peripheral neurons, but no evidence suggests that such coupling occurs in central neurons during the firing of action potentials. Using fast Ca2+ imaging in CA1 neurons from hippocampal slices, we found evidence for CICR during action potential-evoked Ca2+ transients. A low concentration of caffeine enhanced Ca2+ transient amplitude, whereas a higher concentration reduced it. Simultaneous Ca2+ imaging and whole-cell recordings showed that membrane potential, action potential amplitude, and waveform were unchanged during caffeine application. The enhancement of Ca2+ transients by caffeine was not affected by the L-type channel blocker nifedipine, the phosphodiesterase inhibitor IBMX, the adenylyl cyclase activator forskolin, or the PKA antagonist H-89. However, thapsigargin or ryanodine, which both empty intracellular Ca2+ stores, occluded this effect. In addition, thapsigargin, ryanodine, and cyclopiazonic acid reduced action potential-evoked Ca2+ transients in the absence of caffeine. These results suggest that Ca2+ release from ryanodine-sensitive stores contributes to Ca2+ signals triggered by action potentials in CA1 neurons.

The neuronal nicotinic synapse in tissue slices of the adrenal medulla was studied with whole-cell patch-clamp. Excitatory postsynaptic currents (EPSCs) were evoked by local field stimulation or occurred spontaneously especially when external [K+l was increased. EPSCs were carried by channels sharing biophysical and pharmacological properties of neuronal-type nicotinic receptors (nAChRs). A single-channel conductance (y) of 43-45 pS was found from nonstationary variance analysis of EPSCs. Spontaneous EPSCs were tetrodotoxin-insensitive and Ca2+-dependent and occurred in burst-like clusters. Quantal analysis of spontaneous EPSCs gave a quantal size of 20 pA and amplitude histograms were well described by binomial models with low values of quantal content, consistent with a small number of spontaneously active release sites. However, rare large amplitude EPSCs suggest that the total number of sites is higher and that extrajunctional receptors are involved. Our estimates of quantal content and size at the chromaffin cell neuronal nicotinic synapse may be useful in characterizing central neuronal-type nicotinic receptor-mediated cholinergic synaptic transmission.The neuronal nicotinic receptor (nAChR) family displays a large diversity of a and ,B subunit composition and, correspondingly, differences in pharmacological profile, singlechannel conductance, and distribution in the central nervous system and autonomic ganglia (1, 2). Activation of central somatic nAChRs either depolarizes neurons from prefrontal cortex (3), medial habenula (4), and locus coeruleus (5) or hyperpolarizes neurons from dorsolateral septal nucleus (6). As well, nAChRs appear to be involved in presynaptic control of transmitter release (7,8). However, apart from the early discovery of nicotinic excitatory synaptic potentials in cat Renshaw cells (9), postsynaptic nAChRs have not been shown to be directly involved in central synaptic transmission. Thus, neuronal nicotinic synapses have been mostly studied in vertebrate autonomic ganglia (10-13) and recently in the guineapig adrenal gland (14, 15). Acetylcholine (ACh) release has been shown to be quantal at ganglionic synapses (16), but detailed quantal analysis was not done due to the poor resolution associated with intracellular recording. Detection of the postsynaptic action of a single quantum is a prerequisite to quantal analysis and determination of quantal parameters (17), thereby allowing study of pre-and postsynaptic factors involved in synaptic transmission (18,19).Cholinergic synapses arising from splanchnic nerve terminals are present on chromaffin cells (20). These cells are small, rounded, electrically compact and well suited for patch-clamp studies. ACh-induced current in chromaffin cells is due to the activation of a3 subunit containing nAChRs (21,22), which are also expressed in the hippocampus, medial habenula, lateral geniculate, and granular layer of cerebellum (2, 23). The (32 and perhaps also the 134 subunit appear to be coexpressed in chromaffin cells, given the se...

1. Whole-cell voltage-activated currents from single zona fasciculata (ZF) cells from rat adrenal glands were studied. T-and L-type Ca2+ currents and a slowly inactivating A-type K+ current were the three major currents observed.