Endogenous activation of adenosine A1 receptors, but not P2X receptors, during high-frequency synaptic transmission at the calyx of Held

Activation of presynaptic receptors plays an important role in modulation of transmission at many synapses, particularly during high-frequency trains of stimulation. Adenosine-triphosphate (ATP) is coreleased with several neurotransmitters and acts at presynaptic sites to reduce transmitter release; such presynaptic P2X receptors occur at inhibitory and excitatory terminals in the medial nucleus of the trapezoid body (MNTB). We have investigated the mechanism of purinergic modulation during high-frequency repetitive stimulation at the calyx of Held synapse. Suppression of calyceal excitatory postsynaptic currents (EPSCs) by ATP and ATPgammaS (100 microM) was mimicked by adenosine application and was blocked by DPCPX (10 microM), indicating mediation by adenosine A1 receptors. DPCPX enhanced EPSC amplitudes during high-frequency synaptic stimulation, suggesting that adenosine has a physiological role in modulating transmission at the calyx. The Luciferin-Luciferase method was used to probe for endogenous ATP release (at 37 degrees C), but no release was detected. Blockers of ectonucleotidases also had no effect on endogenous synaptic depression, suggesting that it is adenosine acting on A1 receptors, rather than degradation of released ATP, which accounts for presynaptic purinergic suppression of synaptic transmission during physiological stimulus trains at this glutamatergic synapse.     

The pool of fast releasing vesicles is augmented by myosin light chain kinase inhibition at the calyx of Held synapse

Synaptic strength is determined by release probability and the size of the readily releasable pool of docked vesicles. Here we describe the effects of blocking myosin light chain kinase (MLCK), a cytoskeletal regulatory protein thought to be involved in myosin-mediated vesicle transport, on synaptic transmission at the mouse calyx of Held synapse. Application of three different MLCK inhibitors increased the amplitude of the early excitatory postsynaptic currents (EPSCs) in a stimulus train, without affecting the late steady-state EPSCs. A presynaptic locus of action for MLCK inhibitors was confirmed by an increase in the frequency of miniature EPSCs that left their average amplitude unchanged. MLCK inhibition did not affect presynaptic Ca²⁺ currents or action potential waveform. Moreover, Ca²⁺ imaging experiments showed that [Ca²⁺]_i transients elicited by 100-Hz stimulus trains were not altered by MLCK inhibition. Studies using high-frequency stimulus trains indicated that MLCK inhibitors increase vesicle pool size, but do not significantly alter release probability. Accordingly, when AMPA-receptor desensitization was minimized, EPSC paired-pulse ratios were unaltered by MLCK inhibition, suggesting that release probability remains unaltered. MLCK inhibition potentiated EPSCs even when presynaptic Ca²⁺ buffering was greatly enhanced by treating slices with EGTA-AM. In addition, MLCK inhibition did not affect the rate of recovery from short-term depression. Finally, developmental studies revealed that EPSC potentiation by MLCK inhibition starts at postnatal day 5 (P5) and remains strong during synaptic maturation up to P18. Overall, our data suggest that MLCK plays a crucial role in determining the size of the pool of synaptic vesicles that undergo fast release at a CNS synapse.     

Frequency-dependent synaptic depression and the balance of excitation and inhibition in the neocortex

The stability of cortical neuron activity in vivo suggests that the firing rates of both excitatory and inhibitory neurons are dynamically adjusted. Using dual recordings from excitatory pyramidal neurons and inhibitory fast-spiking neurons in neocortical slices, we report that sustained activation by trains of several hundred presynaptic spikes resulted in much stronger depression of synaptic currents at excitatory synapses than at inhibitory ones. The steady-state synaptic depression was frequency dependent and reflected presynaptic function. These results suggest that inhibitory terminals of fast-spiking cells are better equipped to support prolonged transmitter release at a high frequency compared with excitatory ones. This difference in frequency-dependent depression could produce a relative increase in the impact of inhibition during periods of high global activity and promote the stability of cortical circuits.     

Short-term synaptic depression in the neonatal mouse spinal cord: effects of calcium and temperature

We have studied short-term synaptic depression of excitatory postsynaptic potentials (EPSPs) in lumbosacral motoneurons in the isolated, in vitro spinal cord of neonatal mice at 2-4 days postnatal age. We used 2-amino-5-phosphonovaleric acid (AP5; 100 microM) to suppress spontaneous and stimulus-evoked polysynaptic activity. Monosynaptic EPSPs were generated by trains of 10 pulses stimuli delivered to a dorsal root at eight frequencies between 0.125 and 16 Hz. The amplitudes of the second (R2), third (R3), and the average of R8, R9, and R10 (tail) EPSPs, normalized by the first EPSP (R1), defined the shapes of synaptic depression curves. Tail responses were increasingly depressed as stimulation frequency increased but R2 and R3 exhibited relative facilitation at frequencies >1 Hz. Control experiments indicated that the depression curves were not explained by presynaptic activation failure. Lowering external Ca(2+) concentration ([Ca(2+)](o)) from 2.0 to 0.8 mM without changing [Mg(2+)](o) reduced average R1 amplitudes and R2 depression with little change in tail depression. Conversely, increasing [Ca(2+)](o) to 4.0 mM increased average R1 amplitude and R2 depression but again did not change tail depression. Increasing the bath temperature from 24 to 32 degrees C produced little change in R1 amplitudes but markedly reduced the depression of all responses at most frequencies. We developed an empirical model, based on mechanisms described in more accessible synaptic systems, that assumes: transmitter is released from a constant fraction, f, of release-ready elements in two presynaptic compartments (N and S) that are subsequently renewed by independent processes with exponential time constants (tau(N) and tau(S)); an activation-dependent facilitation of transmitter release with constant increment and fast exponential decay; and a more slowly decaying, activation-dependent augmentation of the rate of renewal (tau(N)) of N. The model gave satisfactory fits to data from all [Ca(2+)](o) conditions and implied that f and the increments of the facilitation and augmentation processes were all changed in the same direction as [Ca(2+)](o), without changing the time constants. In contrast, model fits to the 32 degrees C data implied that the process time constants all decreased by 40-45% while the presumably Ca(2+)-related weighting factors were unchanged. The model also successfully matched the normalized amplitudes of EPSPs during trains with irregular intervals.     

Simulation of synaptic depression, posttetanic potentiation, and presynaptic facilitation of synaptic potentials from sensory neurons mediating gill-withdrawal reflex in Aplysia

The defensive gill-withdrawal reflex in Aplysia has proven to be an attractive system for analyzing the neural mechanisms underlying simple forms of learning such as habituation, sensitization, and classic conditioning. Previous studies have shown that habituation is associated with synaptic depression and sensitization with presynaptic facilitation of transmitter release from sensory neurons mediating the reflex. The synaptic depression, in turn, is associated with a decrease in Ca2+ currents in the sensory neurons, whereas presynaptic facilitation with increased Ca2+ currents produced indirectly by a decrease in a novel serotonergic sensitive K+ current. The present work represents an initial quantitative examination of the extent to which these mechanisms account for each of these types of synaptic plasticity. To address these issues a lumped parameter mathematical model of the sensory neuron release process was constructed. Major components of this model include Ca2+-channel inactivation, Ca2+-mediated neurotransmitter release and mobilization, and readily releasable and upstream feeding pools of neurotransmitter. In the model, release of neurotransmitter has a linear function of Ca2+ concentration and is not affected directly by residual Ca2+. The model not only simulates the data of synaptic depression and recovery from depression, but also qualitatively predicts other features of neurotransmitter release that it was not designed to fit. These include features of synaptic depression with high and low levels of transmitter release, posttetanic potentiation, a steep relationship between action potential duration and transmitter release, enhanced release produced by broadening the sensory neuron action potential (presynaptic facilitation), and dramatic synaptic depression with two closely spaced tetraethylammonium (TEA) spikes. The model cannot account fully for synaptic depression with empirically observed somatic Ca2+-current kinetics. Rather a large component of synaptic depression is due to reduction to the pools of releasable neurotransmitter (depletion). In the model when spike durations are greater than 15-20 ms, spike broadening produces little facilitation. However, when spike durations are more physiological, spike broadening leads to enhanced transmitter release.     

Relative roles of different mechanisms of depression at the mouse endbulb of Held

Several mechanisms can underlie short-term synaptic depression, including vesicle depletion, receptor desensitization, and changes in presynaptic release probability. To determine which mechanisms affect depression under physiological conditions, we studied the synapse formed by auditory nerve fibers onto bushy cells in the anteroventral cochlear nucleus (the "endbulb of Held") using voltage-clamp recordings of brain slices from P15-P21 mice near physiological temperatures. Depression of both alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) excitatory postsynaptic currents (EPSCs) showed two phases of recovery. The fast component of depression for the AMPA EPSC was eliminated by cyclothiazide and aniracetam, suggesting it results from desensitization. The fast component of depression for the NMDA EPSC was reduced by the low-affinity antagonist l-AP5, suggesting it results from saturation. The remaining depression in AMPA and NMDA components is identical and therefore presynaptic in origin. It is likely to result from presynaptic vesicle depletion. Recovery from depression after trains of activity was slowed by the application of EGTA-AM, suggesting that the endbulb has a residual-calcium-dependent form of recovery. We developed a model that incorporates depletion, desensitization, and calcium-dependent recovery. This model replicated experimental findings over a range of experimental conditions. The model further indicated that desensitization plays only a minor role during prolonged activity, in large part because presynaptic release is so depleted. Thus depletion appears to be the dominant mechanism of depression at the endbulb during normal activity. Furthermore, calcium-dependent recovery at the endbulb is critical to prevent complete rundown during high activity and to preserve the reliability of information transmission.     

Synaptic depression in visual cortex tissue slices: an in vitro model for cortical neuron adaptation

Synaptic depression was assessed from intracellular recordings in cortical tissue slices. Evoked postsynaptic potentials exhibited synaptic depression with an exponential or double exponential decrease (time constants: <1–30 s) in amplitude during repetitive afferent stimulation by short trains of suprathreshold stimuli. Depressed synaptic responses recovered with an exponential time course (time constants: 10 s-8 min) during presentation of similar short trains of stimuli every 5 or 10 s. Cortical cells recorded extracellularly in cat visual cortex show similar time constants of response decrement during adaptation to moving stripes. Postsynaptic voltage- or ion-regulated conductances and chloride conductances do not appear to be involved in synaptic depression. Input resistance changes and effects of injection of chloride indicate a lack of GABA_A receptor-mediated effects. Hyperpolarizing or depolarizing neurons, and pairing polarization with afferent stimulation, also did not affect synaptic depression. This distinguishes these processes from long-term depression and long-term potentiation. Our results suggest that the most likely mechanisms of synaptic depression and adaptation in cortical cells are presynaptic decrease in transmitter release and/or receptor desensitization. Short-term postsynaptic changes may also occur after synaptic depression.     

Developmental profiles of glutamate receptors and synaptic transmission at a single synapse in the mouse auditory brainstem

Using whole-cell recordings from presynaptic terminals and postsynaptic principal neurons in the mouse medial nucleus of the trapezoid body (MNTB), we have characterized properties of the calyx of Held synapse during the first three postnatal weeks. We observed that evoked excitatory postsynaptic currents (EPSCs) mediated by NMDA receptors (NMDAR) increased until postnatal day 11/12 (P11/12) after which they declined to very low or undetectable levels at P16. Meanwhile, EPSCs mediated by AMPA receptors (AMPAR) showed an approximate three-fold increase in amplitude. These changes were paralleled by NMDAR and AMPAR currents evoked by exogenous NMDA and kainate to MNTB neurons except that whole-cell kainate currents remained constant after P7/8 while AMPAR-EPSCs continued to increase. We found that the decay time constant τ for NMDAR-EPSCs and AMPAR-EPSCs declined by about 30 % and 70 %, respectively. Analyses of NMDAR-EPSCs with subunit-specific pharmacological agents including ifenprodil, N,N,N',N' -tetrakis(2-pyridylmethyl)-ethylenediamine (TPEN), zinc and Mg²⁺ revealed subtle developmental changes in subunit composition. As maturation progressed, this synapse displayed a reduction in the number of presynaptic spike failures and the extent of synaptic depression in response to trains of stimuli (50–300 Hz) while the recovery rate from depression accelerated. These results demonstrate profound changes in the size and kinetics of postsynaptic glutamate receptors and in the spike-firing capability of presynaptic terminals at the calyx of Held-MNTB synapse during early development. We suggest that these concurrent presynaptic and postsynaptic adaptations represent important steps for synapse consolidation and refinement and ultimately for the development of fast high-fidelity transmission at this synapse.     

Coincident glutamatergic and cholinergic inputs transiently depress glutamate release at rat schaffer collateral synapses

The mammalian hippocampus, together with subcortical and cortical areas, is responsible for some forms of learning and memory. Proper hippocampal function depends on the highly dynamic nature of its circuitry, including the ability of synapses to change their strength for brief to long periods of time. In this study, we focused on a transient depression of glutamatergic synaptic transmission at Schaffer collateral synapses in acute hippocampal slices. The depression of evoked excitatory postsynaptic current (EPSC) amplitudes, herein called transient depression, follows brief trains of synaptic stimulation in stratum radiatum of CA1 and lasts for 2-3 min. Depression results from a decrease in presynaptic glutamate release, as NMDA-receptor-mediated EPSCs and composite EPSCs are depressed similarly and depression is accompanied by an increase in the paired-pulse ratio. Transient depression is prevented by blockade of metabotropic glutamate and acetylcholine receptors, presumably located presynaptically. These two receptor types--acting together--cause depression. Blockade of a single receptor type necessitates significantly stronger conditioning trains for triggering depression. Addition of an acetylcholinesterase inhibitor enables depression from previously insufficient conditioning trains. Furthermore, a strong coincident, but not causal, relationship existed between presynaptic depression and postsynaptic internal Ca(2+) release, emphasizing the potential importance of functional interactions between presynaptic and postsynaptic effects of convergent cholinergic and glutamatergic inputs to CA1. These convergent afferents, one intrinsic to the hippocampus and the other likely originating in the medial septum, may regulate CA1 network activity, the induction of long-term synaptic plasticity, and ultimately hippocampal function.     

Presynaptic plasticity at two giant auditory synapses in normal and deaf mice

Large calyceal synapses are often regarded as simple relay points, built for high-fidelity and high-frequency synaptic transmission and a minimal requirement for synaptic plasticity, but this view is oversimplified. Calyceal synapses can exhibit surprising activity-dependent developmental plasticity. Here we compare basal synaptic transmission and activity-dependent plasticity at two stereotypical calyceal synapses in the auditory pathway, the endbulb and the calyx of Held. Basal synaptic transmission was more powerful at the calyx than the endbulb synapse: the amplitude of evoked AMPA receptor-mediated excitatory postsynaptic currents (eEPSCs) was significantly greater at the calyx, as were the release probability, and the number of release sites. The quantal amplitude was smaller at the calyx, consistent with the smaller amplitude of spontaneous miniature EPSCs at this synapse. High-frequency trains of stimuli revealed that the calyx had a larger readily releasable pool of vesicles (RRP), less tetanic depression and less asynchronous transmitter release. Activity-dependent synaptic plasticity was assessed in congenitally deaf mutant mice ( dn/dn ). Previously we showed that a lack of synaptic activity in deaf mice increases synaptic strength at the endbulb of Held via presynaptic mechanisms. In contrast, we have now found that deafness does not affect synaptic transmission at the calyx synapse, as eEPSC and mEPSC amplitude, release probability, number of release sites, size of RRP, tetanic depression and asynchronous release were unchanged compared to normal mice. Synaptic transmission at the calyx synapse is more powerful and has less capacity for developmental plasticity compared to the endbulb synapse.     

Expression of the P/Q (Cav2.1) calcium channel in nodose sensory neurons and arterial baroreceptors

The predominant calcium current in nodose sensory neurons, including the subpopulation of baroreceptor neurons, is the N-type channel, Cav2.2. It is also the primary calcium channel responsible for transmitter release at their presynaptic terminals in the nucleus of the solitary tract in the brainstem. The P/Q channel, Cav2.1, the other major calcium channel responsible for transmitter release at mammalian synapses, represents only 15–20% of total calcium current in the general population of sensory neurons and makes a minor contribution to transmitter release at the presynaptic terminal. In the present study we identified a subpopulation of the largest nodose neurons (capacitance > 50 pF) in which, surprisingly, Cav2.1 represents over 50% of the total calcium current, differing from the remainder of the population. Consistent with these electrophysiological data, anti-Cav2.1 antibody labeling was more membrane delimited in a subgroup of the large neurons in slices of nodose ganglia. Data reported in other synapses in the central nervous system assign different roles in synaptic information transfer to the P/Q-type versus N-type calcium channels. The study raises the possibility that the P/Q channel which has been associated with high fidelity transmission at other central synapses serves a similar function in this group of large myelinated sensory afferents, including arterial baroreceptors where a high frequency regular discharge pattern signals the pressure pulse. This contrasts to the irregular lower frequency discharge of the unmyelinated fibers that make up the majority of the sensory population and that utilize the N-type channel in synaptic transmission.     

Depression and recovery of transmission at the squid giant synapse.

1. The process of synaptic depression and recovery were studied in the squid (Loligo pealii) giant synapse with intracellular recording and stimulating electrodes in the prescence of tetrodotoxin (10-minus 7 M). 2. When the synapse was stimulated at 50 Hz, depression occurred rapidly. Recovery after the tetanus was a first-order process with an average recovery time constant of 4-9 sec. The rate of recovery was independent of the amplitude of the post-synaptic potential (p.s.p.) or the degree of depression. 3. For the first five to seven p.s.p.s in the train there was a linear relationship between depression and the total amount of transmitter previously released. This may indicate that depression in this preparation was caused by the depletion of the presynaptic store of transmitter (S). 4. Assuming that this interpretation was correct, we could show that recovery from depression during the tetanus (i.e. 'mobilization') proceeded about 10 times faster than after the end of the tetanus. 5. When the amplitude of the p.s.p. was varied by changing the bathing calcium concentration, [Ca], the degree of depression was correlated to the amplitude of the p.s.p. 6. When the amplitude of the p.s.p. was increased by increasing pre-synaptic depolarization, synaptic depression was found to increase as well. However, synaptic depression increased less than the amplitude of the p.s.p., the relationship between these two measures being non-linear. 7. This finding is interpreted to indicate that the transmitter stores, S, are closely related to the area of the presynaptic membrane which is sufficiently depolarized to release transmitter.     

Contribution of presynaptic Na(+) channel inactivation to paired-pulse synaptic depression in cultured hippocampal neurons

Paired-pulse depression (PPD) of synaptic transmission is important for neuronal information processing. Historically, depletion of the readily releasable pool of synaptic vesicles has been proposed as the major component of PPD. Recent results suggest, however, that other mechanisms may be involved in PPD, including inactivation of presynaptic voltage-dependent sodium channels (NaChs), which may influence coupling of action potentials to transmitter release. In hippocampal cultures, we have examined the potential role and relative contribution of presynaptic NaCh inactivation in excitatory postsynaptic current (EPSC) PPD. Based on current- and voltage-clamp recordings from somas, our data suggest that NaCh inactivation could potentially participate in PPD. Paired stimulation of somatic action potentials (20- to 100-ms interval) results in subtle changes in action potential shape that are mimicked by low concentrations of tetrodotoxin (TTX) and that appear to be generated by a combination of fast and slow recovery from NaCh inactivation. Dilute concentrations of TTX dramatically depress glutamate release. However, we find evidence for only minimal contribution of NaCh inactivation to EPSC PPD under basal conditions. Hyperpolarization of presynaptic elements to speed recovery from inactivation or increasing the driving force on Na(+) ions through active NaChs had minimal effects on PPD while more robustly reversing the effects of pharmacological NaCh blockade. On the other hand, slight depolarization of the presynaptic membrane potential, by elevating extracellular [K(+)](o), significantly increased PPD and frequency-dependent depression of EPSCs during short trains of action potentials. The results suggest that NaCh inactivation is poised to modulate EPSC amplitude with small tonic depolarizations that likely occur with physiological or pathophysiological activity.     

Long-term potentiation in the hippocampus

Long-term potentiation of field and single neuronal responses recorded in various hippocampal fields is described on the basis of author's and literary data. Most of intrahippocampal and extrinsic connections in both in vivo and in vitro hippocampal preparations show this phenomenon after one or several conditioning trains of comparatively short duration (20 s or less) at various frequencies (from 10 to 400 Hz). Properties of hippocampal potentiation are described. The properties include long term persistence (hours and days) of the potentiated response, its low frequency depression, self-restoration after the depression, specificity of the potentiation for the tetanized pathway, necessity of activation of a sufficient number of neuronal elements (‘cooperativity’) to produce the potentiation, possible involvement of ‘reinforcing’ brain structures during conditioning tetanization. These properties are distinct from those of ‘usual’ short-term post-tetanic potentiation and lead to the suggestion that the neuronal mechanisms underlying long-term potentiation are similar to those underlying memory and behavioralconditioned reflex. Neurophysiological mechanisms of long-term potentiation are discussed. The main mechanism consists in an increase in efficacy of excitatory synapses as shown by various methods including intracellular recording and quantal analysis. The latter favours presynaptic localization of the changes of synaptic efficacy showing increase in the number of transmitter quanta released per presynaptic impulse. However, changes in the number of subsynaptic receptors or localized changes in dendritic postsynaptic membrane are not excluded. Biochemical studies indicate the increase in transmitter release and calcium-dependent phosphorylation of pyruvate dehydrogenase after tetanization. Instances of persistent response facilitations at other levels of the vertebrate central nervous system (especially at neocortical level) are considered and compared with hippocampal long-term potentiation.

It is suggested that modifiable excitatory synapses necessary for learning have been identified in studies of long-term potentiation. These synapses are presumably modified as a result of close sequential activation of the following three structures: excitatory presynaptic fibers, the postsynaptic neuron and a ‘reinforcing’ brain system.     

Visualization of changes in presynaptic function during long-term synaptic plasticity.

Controversy exists regarding the site of modification of synaptic transmission during long-term plasticity in the mammalian hippocampus. Here we used a fluorescent marker of presynaptic activity, FM 1-43, to directly image changes in presynaptic function during both short-term and long-term forms of plasticity at presynaptic boutons of CA3-CA1 excitatory synapses in acute hippocampal slices. We demonstrated enhanced presynaptic function during long-term potentiation (LTP) induced either chemically (with tetraethylammonium), or by high-frequency (200-Hz) electrical stimulation. Both of these forms of LTP required activation of L-type voltage-gated calcium channels and NMDA receptors in the postsynaptic CA1 neuron. These results thus implied that a long-lasting increase in the efficacy of synaptic transmission is likely to depend, at least in part, on enhanced transmitter release from the presynaptic neuron.     

Synaptic depression related to presynaptic axon conduction block.

1. The depression of synaptic transmission, which occurs during prolonged repetitive activation, was examined in the opener muscle of the crayfish walking leg. 2. Excitatory post-synaptic potentials (e.p.s.p.s) initially facilitated but then declined to low amplitudes after about 4000 stimulus pulses had been delivered; this depression is presynaptic in origin; 3. Axon conduction blocks occured at points of bifurcation along the entire length of the presynaptic nerve. This resulted in failure of the nerve impulse to invade some branches of the terminal arborization. 4. Nerve terminal invasion failure caused either intermittent or complete inactiviation of some synaptic release sites; this was associated with depression of the post-synaptic response. 5. The statistics of transmitter release during prolonged repetitive stimulation were examined by focal extracellular recording methods. Transmitter release could be described by binomial statistics, and depression involved a drop in m, n and p. 6. The rate of spontaneous quantal release did not decrease, however, arguing against transmitter depletion. 7. It is concluded that repetitive stimulation eventually leads to depolarization of the axon membrane. This causes impulse propagation failure which reduces the number of synaptic release sites that are activated and mimics a drop in the effective stimulation rate; both effects cause synaptic depression.     

Synaptic transfer at a vertebrate central nervous system synapse.

1. The relation between presynaptic depolarization and transmitter release was examined at a synapse between a Müller axon and a lateral interneurone in the spinal cord of the lamprey. Two micro-electrodes, one for passing current and the other for recording the resulting voltage change, were placed in the presynaptic axon; a single electrode for recording the post-synaptic potential produced by release of transmitter was placed in the post-synaptic cell. 2. When action potentials were blocked with tetrodotoxin, brief depolarizing pulses in the presynaptic fibre were as effective as the action potential had been in producing transmitter release. 3. The release process had an apparent threshold depolarization of 40-50 mV and saturated at presynaptic depolarizations of the order of 100 mV. Increasing the duration of the presynaptic pulse increased the maximum level of release. 4. Displacing the presynaptic voltage recording electrode from the position of synaptic contact toward the current passing electrode increased the apparent depolarization required to produce a given level of transmitter release. This shift in the input-output relation was consistent in magnitude with the voltage attenuation between the presynaptic recording electrode and the synapse expected from the space constant of the fibre. 5. The effect of conditioning hyperpolarization and depolarization of the presynaptic fibre on subsequent transmitter release by brief depolarizing pulses was examined. No effect was observed when the presynaptic recording electrode was in the region of synaptic contact. When the presynaptic electrode was not so positioned, conditioning effects were observed which depended on electode position and could be attributed to changes in the space constant of the presynaptic fibre. No conditioning effects were observed on transmitter release by the action potential.     

Release kinetics, quantal parameters and their modulation during short-term depression at a developing synapse in the rat CNS

We have characterized developmental changes in the kinetics and quantal parameters of action potential (AP)-evoked neurotransmitter release during maturation of the calyx of Held synapse. Quantal size ( q ) and peak amplitudes of evoked EPSCs increased moderately, whereas the fraction of vesicles released by single APs decreased. During synaptic depression induced in postnatal day (P) 5–7 synapses by 10–100 Hz stimulation, q declined rapidly to 40–12% of its initial value. The decrease in q was generally smaller in more mature synapses (P12–14), but quite severe for frequencies ≥ 300 Hz. The stronger decline of q in immature synapses resulted from a slower recovery from desensitization, presumably due to delayed glutamate clearance. Recovery from this desensitization followed an exponential time course with a time constant of ∼480 ms in P5–7 synapses, and sped up > 20-fold during maturation. Deconvolution analysis of EPSCs revealed a significant acceleration of the release time course during development, which was accompanied by a 2-fold increase of the peak release rate. During long 100 Hz trains, more mature synapses were able to sustain average rates of 8–10 quanta s⁻¹ per active zone for phasic release. The rates of asynchronous vesicle release increased transiently > 35-fold immediately after such stimuli and decayed rapidly with an exponential time constant of ∼50 ms to low resting levels of spontaneous release. However, even following extended periods of 100 Hz stimulation, the amount of asynchronous release was relatively minor with peak rates of less than 5% of the average rate of synchronous release measured at steady state during the tetani. Therefore, a multitude of mechanisms seems to converge on the generation of fast, temporally precise and reliable high-frequency transmission at the mature calyx of Held synapse.     

Short-term synaptic depression and recovery at the mature mammalian endbulb of Held synapse in mice

The endbulb of Held synapses between the auditory nerve fibers (ANF) and cochlear nucleus bushy neurons convey fine temporal information embedded in the incoming acoustic signal. The dynamics of synaptic depression and recovery is a key in regulating synaptic transmission at the endbulb synapse. We studied short-term synaptic depression and recovery in mature (P22-38) CBA mice with stimulation rates that were comparable to sound-driven activities recorded in vivo. Synaptic depression in mature mice is less severe ( approximately 40% at 100 Hz) than reported for immature animals and the depression is predominately due to depletion of releasable vesicles. Recovery from depression depends on the rate of activity and accumulation of intracellular Ca(2+) at the presynaptic terminal. With a regular stimulus train at 100 Hz in 2 mM external [Ca(2+)], the recovery from depletion was slow (tau(slow), approximately 2 s). In contrast, a fast (tau(fast), approximately 25 ms), Ca(2+)-dependent recovery followed by a slower recovery (tau(slow), approximately 2 s) was seen when stimulus rates or external [Ca(2+)] increased. In normal [Ca(2+)], recovery from a 100-Hz Poisson-like train is rapid, suggesting that Poisson-like trains produce a higher internal [Ca(2+)] than regular trains. Moreover, the fast recovery was slowed by approximately twofold in the presence of calmidazolium, a Ca(2+)/calmodulin inhibitor. Our results suggest that endbulb synapses from high spontaneous firing rate auditory nerve fibers normally operate in a depressed state. The accelerated synaptic recovery during high rates of activity is likely to ensure that reliable synaptic transmission can be achieved at the endbulb synapse.