Unipolar brush cells form a glutamatergic projection system within the mouse cerebellar cortex.

Unipolar brush cells (UBCs) of the mammalian vestibulocerebellum receive mossy fiber projections primarily from the vestibular ganglion and vestibular nuclei. Recently, the axons of UBCs have been shown to generate an extensive system of cortex-intrinsic mossy fibers, which resemble traditional cerebellar mossy fiber afferents and synapse with granule cell dendrites and other UBCs. However, the neurotransmitter used by the UBC axon is still unknown. In this study, we used long-term organotypic slice cultures of the isolated nodulus (lobule X) from postnatal day 8 mouse cerebella to identify the neurotransmitter and receptors at synapses of the UBC axon terminals, relying on the notion that, in these cultures, all of the cortex-extrinsic fibers had degenerated during the first few days in vitro. Quantification of glutamate immunogold labeling showed that the UBC axon terminals have the same high gold-particle density as the glutamatergic parallel fiber varicosities. Furthermore, UBCs identified by calretinin immunoreactivity expressed the glutamate receptor subunits GluR2/3, NMDAR1, and mGluR2/3, like they do in the mature mouse cerebellum in situ. Evoked excitatory postsynaptic currents (EPSCs), spontaneous EPSCs, and burst discharges were demonstrated in UBCs and granule cells by patch-clamp recording. Both the evoked and spontaneous EPSCs were blocked by ionotropic glutamate receptor antagonists CNQX and D-AP5. We conclude that neurotransmission at the UBC axon terminals is glutamatergic. Thus, UBCs provide a powerful network of feedforward excitation within the granular layer, which may amplify vestibular signals and synchronize activity in clusters of functionally related granule cells which project vertically to patches of Purkinje cells.

Desensitization of mutant acetylcholine receptors in transgenic mice reduces the amplitude of neuromuscular synaptic currents.

While the slow onset of desensitization of nicotinic acetylcholine receptors (AChRs), relative to the rate of acetylcholine removal, excludes this kinetic state from shaping synaptic responses in normal neuromuscular transmission, its role in neuromuscular disorders has not been examined. The slow-channel congenital myasthenic syndrome (SCCMS) is a disorder caused by point mutations in the AChR subunit-encoding genes leading to kinetically abnormal (slow) channels, reduced miniature endplate current amplitudes (MEPCs), and degeneration of the postsynaptic membrane. Because of this complicated picture of kinetic and structural change in the neuromuscular junction, it is difficult to assess the importance of the multiple factors that may be responsible for the reduced endplate current amplitudes, and ultimately the clinical syndrome. In order to address this we have used a transgenic mouse model for the SCCMS that has slow AChR ion channels and reduced endplate responsiveness in the absence of any of the degenerative changes. We found that the reduction in MEPC amplitudes in these mice could not be explained by either reduced AChR number or by reduced AChR channel conductance. Rather, we found that the mutant AChRs in situ manifested an activity-dependent reduction in sensitivity that caused diminished MEPC and endplate current amplitude with nerve stimulation. This observation demonstrates that the basis for the reduction in MEPC amplitudes in the SCCMS may be multifactorial. Moreover, these findings demonstrate that, under conditions that alter their rate of desensitization, the kinetic properties of nicotinic AChRs can control the strength of synaptic responses.

Some effects of d-tubocurarine alone and combined with halothane or isoflurane on neuromuscular transmission.

The mechanisms contributing to the neuromuscular block produced by nondepolarizing muscle relaxants and inhaled anesthetics include: 1) receptor blockade, 2) open or closed ion channel block, 3) decreased transmitter release, and 4) receptor desensitization. In this study we investigated the contributions of receptor and ion channel block. We used the two microelectrode voltage clamp to evaluate miniature end-plate currents (MEPCs) for amplitude and time constant of decay (tau) before and after the application of 1) d-Tubocurarine (DTC) (10(-7)-10(-6) M) alone, and then 2) the same concentration of DTC plus 0.5% or 1.0% halothane or isoflurane delivered by passing compressed air through a flow and temperature compensated vaporizer. The electrodes were maintained in the same cell for the entire experiment. DTC alone decreased MEPC amplitude to 91.3% +/- 4.5% and 65.1% +/- 5.6% of control at 10(-7) and 10(-6) M, respectively. MEPC amplitude with 10(-6) M DTC decreased further to 52.4% +/- 7.6% and 37.4% +/- 7.0% of control after the addition of 0.5% and 1.0% halothane, respectively. After the application of DTC 10(-7) M tau was 94.7% +/- 3.1% of control and decreased to 73.7% +/- 7.1% of control with 10(-6) M DTC. After the application of DTC 10(-6) M the addition of 0.5% and 1% halothane decreased tau to 52.4% +/- 7.6% and 30.0% +/- 5.9% of control. Isoflurane produced similar changes. This study provides evidence that at least some of the augmentation of nondepolarizing relaxants by inhaled anesthetics can be explained by the additive effect of nondepolarizing muscle relaxants and inhaled anesthetics on MEPC amplitude and tau.

Age-induced alteration of neuromuscular transmission: effect of halothane.

Age-induced alteration in neuromuscular transmission and the effect of halothane were examined in 3- and 30-month-old rats with the voltage clamp technique. Significant pre- and post-junctional changes of synaptic transmission occur with advancing age as the frequency of spontaneous release, quantal content and mobilization rate increase and decay time (tau) of miniature end plate current (MEPC) and end-plate current (EPC) is prolonged and altered (bi-exponential). Effects of halothane (0.3-0.88 mM) appeared to be pre-junctional, as the anesthetic decreased the frequency of spontaneous release, end plate current amplitude, quantal content and mobilization rate, had little or no effect on decay time of miniature end plate current and end plate current, and produced no run-down of ionophoretically evoked trains of end plate current. Some of these effects of halothane are more prominent in 30-month-old rats.

Effect of (+/-)-DOI on neuromuscular transmission: a microelectrode study.

DOI (1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane) significantly depressed end plate current (EPC) amplitude. It decreased quantum content, increased the extent of neurally evoked EPC rundown during the train, produced a nonlinear current-voltage relationship, shortened time constant of decay, and depressed iontophoretically evoked EPC. The depressant response of DOI on EPC amplitude was antagonized by 5-HT1-like receptor antagonists, but was resistant to 5-HT2 and 5-HT3 receptor antagonists. This suggests that inhibitory 5-HT receptors roughly correspond to 5-HT1-like receptors.