Voltage-gated calcium channels are well characterized at neuronal somata but less thoroughly understood at the presynaptic terminal where they trigger transmitter release. In order to elucidate how the intrinsic properties of presynaptic calcium channels influence synaptic function, we have made direct recordings of the presynaptic calcium current (I(pCa)) in a brainstem giant synapse called the calyx of Held. The current was pharmacologically classified as P-type and exhibited marked inactivation. The inactivation was largely dependent upon the inward calcium current magnitude rather than the membrane potential, displayed little selectivity between divalent charge carriers (Ca2+, Ba2+ and Sr+), and exhibited slow recovery. Simultaneous pre- and postsynaptic whole-cell recording revealed that I(pCa) inactivation predominantly contributes to posttetanic depression of EPSCs. Thus, because of its slow recovery, I(pCa) inactivation underlies this short-term synaptic plasticity.

Local superfusion of limited dendritic areas with hypertonic or hyperkalemic solutions stimulates the release of quanta from a small population of synapses made on rodent hippocampal neurons maintained in primary culture, and each quantal event can be detected in the postsynaptic neuron. With maintained stimulation, the initial release rate is about 20 quanta per sec per synapse, and this rate declines exponentially to a final low level. These observations can be interpreted as depletion of available quanta and, with this interpretation, a bouton would contain one to two dozen quanta in its readily releasable pool. Tests with a second application of the solution that produces release reveal that the pool of readily releasable quanta is replenished with a time constant of about 10 sec (36°C). The pool of quanta defined in this way may correspond to the population of vesicles docked at the bouton's active zone.Because synapses are so densely packed in brain, the properties of individual, identified boutons are very difficult to investigate. Specifically, no physiological information is available on how many quanta are readily available for release by a single central bouton. Earlier investigators have approached the study of individual central synapses in two ways. First, a giant presynaptic terminal from goldfish bipolar cells has been investigated electrophysiologically with exocytosis and endocytosis detected by measuring changes in membrane capacitance (1, 2). Although this approach has many advantages, a disadvantage is that individual exocytotic events cannot be detected, and the giant terminal contains a large number of specialized release zones that are not typical of most other central synapses. Second, hippocampal synapses have been studied in primary cultures with confocal microscopic techniques that permit rates of exocytosis and endocytosis to be followed optically by the uptake and release of fluorescent dye (3), a method that has been used to address similar issues for the amphibian and mammalian neuromuscular junctions (4,5). This method makes individual boutons available for investigation but has limited temporal and spatial resolution so that single quantal releases cannot be detected.We have exploited the accessibility and low density of synapses made in primary cultures of rodent hippocampal neurons to estimate the number of readily releasable quanta per bouton and the time it takes this pool to be replenished once it is depleted. The method we have employed is to superfuse locally a small, identified population of synapses on a single neuron with a solution that produces exocytosis and to count every miniature excitatory postsynaptic current (mEPSC) that occurs. The high resolution of whole cell recording, together with the relatively low release rates that result when a sufficiently small population of synapses is active, permits us to detect virtually every transmitter quantum released. Two different superfusion solutions ("release solutions") have been employed to produce transmitte...

Metabotropic glutamate receptors (mGluRs) regulate transmitter release at mammalian central synapses. However, because of the difficulty of recording from mammalian presynaptic terminals, the mechanism underlying mGluR-mediated presynaptic inhibition is not known. Here, simultaneous recordings from a giant presynaptic terminal, the calyx of Held, and its postsynaptic target in the medial nucleus of the trapezoid body were obtained in rat brainstem slices. Agonists of mGluRs suppressed a high voltage-activated P/Q-type calcium conductance in the presynaptic terminal, thereby inhibiting transmitter release at this glutamatergic synapse. Because several forms of presynaptic modulation and plasticity are mediated by mGluRs, this identification of a target ion channel is a first step toward elucidation of their molecular mechanism.

P/Q-type presynaptic calcium currents (IpCa) undergo activity-dependent facilitation during repetitive activation at the calyx of the Held synapse. We investigated whether neuronal calcium sensor 1 (NCS-1) may underlie this phenomenon. Direct loading of NCS-1 into the nerve terminal mimicked activity-dependent IpCa facilitation by accelerating the activation time of IpCa in a Ca2+-dependent manner. A presynaptically loaded carboxyl-terminal peptide of NCS-1 abolished IpCa facilitation. These results suggest that residual Ca2+ activates endogenous NCS-1, thereby facilitating IpCa. Because both P/Q-type Ca2+ channels and NCS-1 are widely expressed in mammalian nerve terminals, NCS-1 may contribute to the activity-dependent synaptic facilitation at many synapses.