Synaptic Plasticity in Medial Vestibular Nucleus Neurons: Comparison with Computational Requirements of VOR Adaptation

Background

Vestibulo-ocular reflex (VOR) gain adaptation, a longstanding experimental model of cerebellar learning, utilizes sites of plasticity in both cerebellar cortex and brainstem. However, the mechanisms by which the activity of cortical Purkinje cells may guide synaptic plasticity in brainstem vestibular neurons are unclear. Theoretical analyses indicate that vestibular plasticity should depend upon the correlation between Purkinje cell and vestibular afferent inputs, so that, in gain-down learning for example, increased cortical activity should induce long-term depression (LTD) at vestibular synapses.

Methodology/Principal Findings

Here we expressed this correlational learning rule in its simplest form, as an anti-Hebbian, heterosynaptic spike-timing dependent plasticity interaction between excitatory (vestibular) and inhibitory (floccular) inputs converging on medial vestibular nucleus (MVN) neurons (input-spike-timing dependent plasticity, iSTDP). To test this rule, we stimulated vestibular afferents to evoke EPSCs in rat MVN neurons in vitro. Control EPSC recordings were followed by an induction protocol where membrane hyperpolarizing pulses, mimicking IPSPs evoked by flocculus inputs, were paired with single vestibular nerve stimuli. A robust LTD developed at vestibular synapses when the afferent EPSPs coincided with membrane hyperpolarisation, while EPSPs occurring before or after the simulated IPSPs induced no lasting change. Furthermore, the iSTDP rule also successfully predicted the effects of a complex protocol using EPSP trains designed to mimic classical conditioning.

Conclusions

These results, in strong support of theoretical predictions, suggest that the cerebellum alters the strength of vestibular synapses on MVN neurons through hetero-synaptic, anti-Hebbian iSTDP. Since the iSTDP rule does not depend on post-synaptic firing, it suggests a possible mechanism for VOR adaptation without compromising gaze-holding and VOR performance in vivo.     

Primary neurotransmitters and regulatory substances onto vestibular nucleus neurons.

This review article focused on the primary neurotransmitters involved in transmission from the otolith to the vestibular nucleus (VN), especially in relation to the neurotransmission to the VN neurons (gravity-sensitive neurons) activated by tilt stimulation. The medial vestibular nucleus (MVN) neurons were classified in 8 types (alpha-theta) according to the patterns in response to the clockwise and counterclockwise tilt-stimulations. The tilt-induced firing was inhibited by GDEE (a non-selective glutamate receptor antagonist) and/or atropine (a muscarinic receptor antagonist). Thus, glutamate and/or acetylcholine may serve as the primary neurotransmitters. This conclusion is supported by the previous findings that glutamate exists in the vestibular nerve and is released from the nerve besides the presence of glutamate receptor subtypes in the VN. In addition, acetylcholine induced atropine-reversible firing of MVN neurons, and the enzymes involved in acetylcholine synthesis/metabolism are also found in the VN. Furthermore, serotonin was found to inhibit the MVN neuronal activities via the 5-HT1A receptors. As such, the 5-HT1A agonist, tandospirone, may be effective in preventing and/or treating motion sickness and/or space sickness.     

Intrinsic membrane properties and dynamics of medial vestibular neurons: a simulation

The vestibulo-ocular and vestibulo-spinal network provides the ability to hold gaze fixed on an object during passive head movement. Within that network, most of the second-order neurons of the medial vestibular nucleus (MVNn) compute internal representations of head movement velocity in the horizontal plane. Our previous in vitro studies of the MVNn membrane properties indicated that they may play a major role in determining the dynamic properties of these neurons independently of their connectivity. The present study investigated that hypothesis at a theoretical level. Biophysical models of type A and B MVNn were developed. Two factors were found to be important in modeling tonic and phasic firing activity: the activation of the delayed potassium current and the rate of calcium flux. In addition, the model showed that the strength of the delayed potassium current may determine the different forms of action potentials observed experimentally. These two models (type A and B cells) were examined using depolarizing stimulation, random noise, step, ramp and sinusoidal inputs. For random noise, type A cells showed stable (regular) firing frequencies, while type B cells exhibited irregular activity. With step stimulation, the models exhibited tonic and phasic firing responses, respectively. Using ramp stimulations, frequency versus current curves showed a linear response for the type B neuron model. Finally, with sinusoidal stimulation of increasing frequencies, the type A model demonstrated a decrease in sensitivity, while the type B model exhibited an increase in sensitivity. These theoretical results support the hypothesis that MVNn intrinsic membrane properties specify various types of dynamic properties amongst these cells and therefore contribute to the wide range of dynamic responses which characterize the vestibulo-ocular and vestibulo-spinal network. Received: 1 August 1996 / Accepted in revised form: 16 December 1998     

Low Voltage Activation of KCa1.1 Current by Cav3-KCa1.1 Complexes

Calcium-activated potassium channels of the KCa1.1 class are known to regulate repolarization of action potential discharge through a molecular association with high voltage-activated calcium channels. The current study examined the potential for low voltage-activated Cav3 (T-type) calcium channels to interact with KCa1.1 when expressed in tsA-201 cells and in rat medial vestibular neurons (MVN) in vitro. Expression of the channel α-subunits alone in tsA-201 cells was sufficient to enable Cav3 activation of KCa1.1 current. Cav3 calcium influx induced a 50 mV negative shift in KCa1.1 voltage for activation, an interaction that was blocked by Cav3 or KCa1.1 channel blockers, or high internal EGTA. Cav3 and KCa1.1 channels coimmunoprecipitated from lysates of either tsA-201 cells or rat brain, with Cav3 channels associating with the transmembrane S0 segment of the KCa1.1 N-terminus. KCa1.1 channel activation was closely aligned with Cav3 calcium conductance in that KCa1.1 current shared the same low voltage dependence of Cav3 activation, and was blocked by voltage-dependent inactivation of Cav3 channels or by coexpressing a non calcium-conducting Cav3 channel pore mutant. The Cav3-KCa1.1 interaction was found to function highly effectively in a subset of MVN neurons by activating near –50 mV to contribute to spike repolarization and gain of firing. Modelling data indicate that multiple neighboring Cav3-KCa1.1 complexes must act cooperatively to raise calcium to sufficiently high levels to permit KCa1.1 activation. Together the results identify a novel Cav3-KCa1.1 signaling complex where Cav3-mediated calcium entry enables KCa1.1 activation over a wide range of membrane potentials according to the unique voltage profile of Cav3 calcium channels, greatly extending the roles for KCa1.1 potassium channels in controlling membrane excitability.     

Ethanol potentiates the effect of gamma-aminobutyric acid on medial vestibular nucleus neurons responding to horizontal rotation

Electrophysiological studies were performed to determine whether or not ethanol potentiates the inhibitory effects of gamma-aminobutyric acid (GABA) on medial vestibular nucleus (MVN) neurons responding to horizontal sinusoidal rotation using alpha-chloralose anesthetized cats. The MVN neurons were classified into types I, II, III and IV neurons according to the responses to the horizontal rotation of the animal placed on the turntable in directions ipsilateral and contralateral to the recording site. In addition, the effects of ethanol and GABA on type I neurons were also examined. Micro-osmotic application of ethanol up to 100 nA did not affect the spontaneous firing or the rotation-induced increase in firing of type I neurons. However, the inhibitory effects of GABA up to 50 nA on the rotation-induced increase in firing were potentiated during simultaneous application of ethanol up to 100 nA. This potentiated inhibition was blocked by iontophoretic application of bicuculline (25-150 nA) and picrotoxin (45-150 nA). These results suggest that ethanol potentiates the inhibitory effects of GABA on MVN type I neurons by acting on the GABA receptor and/or receptor-coupled chloride ion channel.     

Inhibition by enkephalin of medial vestibular nucleus neurons responding to horizontal pendular rotation

Electrophysiological studies were performed to determine whether or not enkephalin modulates the activities of medial vestibular nucleus (MVN) neurons responding to horizontal pendular rotation using alpha-chloralose anesthetized cats. The effects of microiontophoretically applied drugs were examined in type I and type II neurons identified according to responses to horizontal, sinusoidal rotation; type I and type II neurons showed an increase and decrease in firing with rotation ipsilateral to the recording site and vice versa with contralateral rotation, respectively. Iontophoretic application of enkephalin suppressed spike firing induced by rotation of the animals in type I neuron, but not in type II neuron. The spike firing induced by iontophoretically applied glutamate was also inhibited during the application of enkephalin. The inhibition by enkephalin of both rotation- and glutamate-induced firing was antagonized by naloxone which was given simultaneously. These results suggest that enkephalin acts on MVN type I neuron to inhibit transmission from the vestibule, thereby controlling vestibulo-ocular reflex.     

A Time Course Analysis of the Electrophysiological Properties of Neurons Differentiated from Human Induced Pluripotent Stem Cells (iPSCs)

Many protocols have been designed to differentiate human embryonic stem cells (ESCs) and human induced pluripotent stem cells (iPSCs) into neurons. Despite the relevance of electrophysiological properties for proper neuronal function, little is known about the evolution over time of important neuronal electrophysiological parameters in iPSC-derived neurons. Yet, understanding the development of basic electrophysiological characteristics of iPSC-derived neurons is critical for evaluating their usefulness in basic and translational research. Therefore, we analyzed the basic electrophysiological parameters of forebrain neurons differentiated from human iPSCs, from day 31 to day 55 after the initiation of neuronal differentiation. We assayed the developmental progression of various properties, including resting membrane potential, action potential, sodium and potassium channel currents, somatic calcium transients and synaptic activity. During the maturation of iPSC-derived neurons, the resting membrane potential became more negative, the expression of voltage-gated sodium channels increased, the membrane became capable of generating action potentials following adequate depolarization and, at day 48–55, 50% of the cells were capable of firing action potentials in response to a prolonged depolarizing current step, of which 30% produced multiple action potentials. The percentage of cells exhibiting miniature excitatory post-synaptic currents increased over time with a significant increase in their frequency and amplitude. These changes were associated with an increase of Ca²⁺ transient frequency. Co-culturing iPSC-derived neurons with mouse glial cells enhanced the development of electrophysiological parameters as compared to pure iPSC-derived neuronal cultures. This study demonstrates the importance of properly evaluating the electrophysiological status of the newly generated neurons when using stem cell technology, as electrophysiological properties of iPSC-derived neurons mature over time.     

Single Versus Repetitive Spiking to the Current Stimulus of A-β Mechanosensitive Neurons in the Crotaline Snake Trigeminal Ganglion

1. Intrasomal recordings of potentials produced by current stimulation in vivo were made from 24 (A-) touch and 19 vibrotactile neurons in the trigeminal ganglion of 29 crotaline snakes, Trimeresurus flavoviridis. 2. Usually touch neurons responded with a single action potential at the beginning of a prolonged depolarizing pulse, whereas all vibrotactile neurons responded with multiple spikes.3. The electrophysiological parameters examined were membrane potential, threshold current, input resistance and capacitance, time constant, rebound latency, and its threshold current. Touch neurons had higher input resistance (and lower input capacitance) than vibrotactile neurons.4. In conclusion, current injection, which elicits a single or multiple spiking, seems a useful way to separate touch neurons from vibrotactile neurons without confirming the receptor response, and some membrane properties are also specific to the sensory modality.     

Unique after-hyperpolarization accompanying action potential inChara globularis

A unique after-hyperpolarization was found in internodal cells ofChara globularis. The cells generated an ordinary action potential due to regenerative depolarization induced by the outward electric current pulse larger than a threshold stimulus. After reaching a depolarizing peak, the membrane potential repolarized and overshooted the resting potential to a value which was somehow 40 mV more negative than the resting potential before stimulation (after-hyperpolarization). Since the membrane resistance increased during the after-hyperpolarization, the after-hyperpolarization is thought to be caused by an increase in the resistance (decrease in the conductance) of the passive diffusion channel.     

Inactivation mode of sodium channels defines the different maximal firing rates of conventional versus atypical midbrain dopamine neurons

Two subpopulations of midbrain dopamine (DA) neurons are known to have different dynamic firing ranges in vitro that correspond to distinct projection targets: the originally identified conventional DA neurons project to the dorsal striatum and the lateral shell of the nucleus accumbens, whereas an atypical DA population with higher maximum firing frequencies projects to prefrontal regions and other limbic regions including the medial shell of nucleus accumbens. Using a computational model, we show that previously identified differences in biophysical properties do not fully account for the larger dynamic range of the atypical population and predict that the major difference is that originally identified conventional cells have larger occupancy of voltage-gated sodium channels in a long-term inactivated state that recovers slowly; stronger sodium and potassium conductances during action potential firing are also predicted for the conventional compared to the atypical DA population. These differences in sodium channel gating imply that longer intervals between spikes are required in the conventional population for full recovery from long-term inactivation induced by the preceding spike, hence the lower maximum frequency. These same differences can also change the bifurcation structure to account for distinct modes of entry into depolarization block: abrupt versus gradual. The model predicted that in cells that have entered depolarization block, it is much more likely that an additional depolarization can evoke an action potential in conventional DA population. New experiments comparing lateral to medial shell projecting neurons confirmed this model prediction, with implications for differential synaptic integration in the two populations.     

A model of the electrophysiological properties of nucleus reticularis thalami neurons.

A model of the electrophysiological properties of rodent nucleus reticularis thalami (NRT) neurons of the dorsal lateral thalamus was developed using Hodgkin-Huxley style equations. The model incorporated voltage-dependent rate constants and kinetics obtained from recent voltage-clamp experiments in vitro. The intrinsic electroresponsivity of the model cell was found to be similar to several empirical observations. Three distinct modes of oscillatory activity were identified: 1) a pattern of slow rhythmic burst firing (0.5-7 Hz) usually associated with membrane potentials negative to approximately -70 mV which resulted from the interplay of ITs and IK(Ca); 2) at membrane potentials from approximately -69 to -62 mV, rhythmic burst firing in the spindle frequency range (7-12 Hz) developed and was immediately followed by a tonic tail of single spike firing after several bursts. The initial bursting rhythm resulted from the interaction of ITs and IK(Ca), with a slow after-depolarization due to ICAN which mediated the later tonic firing; 3) with further depolarization of the membrane potential positive to approximately -61 mV, sustained tonic firing appeared in the 10-200-Hz frequency range depending on the amplitude of the injected current. The frequency of this firing was also dependent on the maximum conductance of the leak current, IK(leak), and an interaction between the fast currents involved in generating action potentials, INa(fast) and IK(DR), and the persistent Na+ current, INa(P). Transitions between different firing modes were identified and studied parametrically.     

Physiological Characterization of Vestibular Efferent Brainstem Neurons Using a Transgenic Mouse Model

The functional role of efferent innervation of the vestibular end-organs in the inner ear remains elusive. This study provides the first physiological characterization of the cholinergic vestibular efferent (VE) neurons in the brainstem by utilizing a transgenic mouse model, expressing eGFP under a choline-acetyltransferase (ChAT)-locus spanning promoter in combination with targeted patch clamp recordings. The intrinsic electrical properties of the eGFP-positive VE neurons were compared to the properties of the lateral olivocochlear (LOC) brainstem neurons, which gives rise to efferent innervation of the cochlea. Both VE and the LOC neurons were marked by their negative resting membrane potential <−75 mV and their passive responses in the hyperpolarizing range. In contrast, the response properties of VE and LOC neurons differed significantly in the depolarizing range. When injected with positive currents, VE neurons fired action potentials faithfully to the onset of depolarization followed by sparse firing with long inter-spike intervals. This response gave rise to a low response gain. The LOC neurons, conversely, responded with a characteristic delayed tonic firing upon depolarizing stimuli, giving rise to higher response gain than the VE neurons. Depolarization triggered large TEA insensitive outward currents with fast inactivation kinetics, indicating A-type potassium currents, in both the inner ear-projecting neuronal types. Immunohistochemistry confirmed expression of Kv4.3 and 4.2 ion channel subunits in both the VE and LOC neurons. The difference in spiking responses to depolarization is related to a two-fold impact of these transient outward currents on somatic integration in the LOC neurons compared to in VE neurons. It is speculated that the physiological properties of the VE neurons might be compatible with a wide-spread control over motion and gravity sensation in the inner ear, providing likewise feed-back amplification of abrupt and strong phasic signals from the semi-circular canals and of tonic signals from the gravito-sensitive macular organs.     

Voltage dependence of subthreshold resonance frequency in layer II of medial entorhinal cortex

The resonance properties of individual neurons in entorhinal cortex (EC) may contribute to their functional properties in awake, behaving rats. Models propose that entorhinal grid cells could arise from shifts in the intrinsic frequency of neurons caused by changes in membrane potential owing to depolarizing input from neurons coding velocity. To test for potential changes in intrinsic frequency, we measured the resonance properties of neurons at different membrane potentials in neurons in medial and lateral EC. In medial entorhinal neurons, the resonant frequency of individual neurons decreased in a linear manner as the membrane potential was depolarized between -70 and -55 mV. At more hyperpolarized membrane potentials, cells asymptotically approached a maximum resonance frequency. Consistent with the previous studies, near resting potential, the cells of the medial EC possessed a decreasing gradient of resonance frequency along the dorsal to ventral axis, and cells of the lateral EC lacked resonant properties, regardless of membrane potential or position along the medial to lateral axis within lateral EC. Application of 10 μM ZD7288, the H-channel blocker, abolished all resonant properties in MEC cells, and resulted in physiological properties very similar to lateral EC cells. These results on resonant properties show a clear change in frequency response with depolarization that could contribute to the generation of grid cell firing properties in the medial EC.     

Altered Neuronal Intrinsic Properties and Reduced Synaptic Transmission of the Rat's Medial Geniculate Body in Salicylate-Induced Tinnitus

Sodium salicylate (NaSal), an aspirin metabolite, can cause tinnitus in animals and human subjects. To explore neural mechanisms underlying salicylate-induced tinnitus, we examined effects of NaSal on neural activities of the medial geniculate body (MGB), an auditory thalamic nucleus that provides the primary and immediate inputs to the auditory cortex, by using the whole-cell patch-clamp recording technique in MGB slices. Rats treated with NaSal (350 mg/kg) showed tinnitus-like behavior as revealed by the gap prepulse inhibition of acoustic startle (GPIAS) paradigm. NaSal (1.4 mM) decreased the membrane input resistance, hyperpolarized the resting membrane potential, suppressed current-evoked firing, changed the action potential, and depressed rebound depolarization in MGB neurons. NaSal also reduced the excitatory and inhibitory postsynaptic response in the MGB evoked by stimulating the brachium of the inferior colliculus. Our results demonstrate that NaSal alters neuronal intrinsic properties and reduces the synaptic transmission of the MGB, which may cause abnormal thalamic outputs to the auditory cortex and contribute to NaSal-induced tinnitus.     

Effects of Weak Environmental Magnetic Fields on the Spontaneous Bioelectrical Activity of Snail Neurons

We examined the effects of 50-Hz magnetic fields in the range of flux densities relevant to our current environmental exposures on action potential (AP), after-hyperpolarization potential (AHP) and neuronal excitability in neurons of land snails, Helix aspersa. It was shown that when the neurons were exposed to magnetic field at the various flux densities, marked changes in neuronal excitability, AP firing frequency and AHP amplitude were seen. These effects seemed to be related to the intensity, type (single and continuous or repeated and cumulative) and length of exposure (18 or 20 min). The extremely low-frequency (ELF) magnetic field exposures affect the excitability of F1 neuronal cells in a nonmonotonic manner, disrupting their normal characteristic and synchronized firing patterns by interfering with the cell membrane electrophysiological properties. Our results could explain one of the mechanisms and sites of action of ELF magnetic fields. A possible explanation of the inhibitory effects of magnetic fields could be a decrease in Ca²⁺ influx through inhibition of voltage-gated Ca²⁺ channels. The detailed mechanism of effect, however, needs to be further studied under voltage-clamp conditions.     

Modifications of Neuronal Activity in the Medial Vestibular Nucleus during Long-Term Vibration

The impulse background activity (BA) of neurons of the rat medial vestibular nucleus (MVN) was subjected to computer analysis, and its modifications related to long-term vibration were studied. It was shown that following 5 days of 2-h-long vibration sessions, statistically significant changes in some basic characteristics of BA generated by MVN neurons were observed. More than a twofold increase in the mean BA frequency and substantial shifts practically of all statistical BA parameters were found after 10 days of vibration. Following a 15-day-long vibration, MVN neurons showed a clear-cut tendency to restore control values of the BA indices, which probably was related to adaptation processes.     

A slow voltage-activated potassium current in rat vagal neurons.

Potassium currents play a key role in controlling the excitability of neurons. In this paper we describe the properties of a novel voltage-activated potassium current in neurons of the rat dorsal motor nucleus of the vagus (DMV). Intracellular recordings were made from DMV neurons in transverse slices of the medulla. Under voltage clamp, depolarization of these neurons from hyperpolarized membrane potentials (more negative than -80 mV) activated two transient outward currents. One had fast kinetics and had properties similar to A-currents. The other current had an activation threshold of around -95 mV (from a holding potential -110 mV) and inactivated with a time constant of about 3s. It had a reversal potential close to the potassium equilibrium potential. This current was not calcium dependent and was not blocked by 4-aminopyridine (5 mM), catechol (5 mM) or tetraethylammonium (20 mM). It was completely inactivated at the resting membrane potential. This current therefore represents a new type of voltage-activated potassium current. It is suggested that this current might act as a brake to repetitive firing when the neuron is depolarized from membrane potentials negative to the resting potential.     

Voltage-Dependent Behavior of Delayed-Firing Neurons in the Rat <Emphasis Type="Italic">Substantia Gelatinosa</Emphasis>

Upon application of a long-lasting rectangular stimulus, neurons of the substantia gelatinosa (SG) display three main types of intrinsic firing behavior, tonic, adapting, and delayed onset. The electrical landmark of delayed-firing neurons (DFNs), i.e., a significant delay before initiation of action potentials (APs), is believed to result from activation of subthreshold A-type K⁺ current (K_A). We checked out this hypothesis by comparing the voltage dependence of the firing delay with steady-state inactivation of K_A in spinal cord slices of 3- to 5-week-old rats. The delay strongly decreased with membrane depolarization and disappeared at ~ –60 mV; herewith the discharge pattern was transformed to either a tonic or an adapting one. This correlated well with inactivation of K_A recorded in a whole-cell mode in low-Cl^– intracellular solution; inactivation was nearly complete at –60 mV (voltage of half-maximum inactivation, V _1/2 ~ –74.5 mV). Unexpectedly, it was found that filling the cells with high-Cl^– solution, to minimize the liquid junction potential, produced at least a 10 mV-difference between voltage dependences of the firing delay and K_A inactivation; the latter shifted toward negativity (V _1/2 ~ –88.3 mV). The results suggest that the K_A and its inactivation properties determine the appearance and voltage dependence of the firing delay in SG neurons; the apparent influence of intracellular Cl^– on inactivation properties needs further investigation.     

Vasopressin induces depolarization and state-dependent firing patterns in rat thalamic paraventricular nucleus neurons in vitro

The thalamic midline paraventricular nucleus (PVT) is prominently innervated by vasopressin-immunoreactive neurons from the suprachiasmatic nucleus (SCN), site of the brain's biological clock. Using patch-clamp recordings in slice preparations taken from Wistar rats during the subjective day, we examined 90 PVT neurons for responses to bath-applied AVP (0.5-2 microM; 1-3 min). In current clamp at resting membrane potentials (-65 +/- 1 mV), PVT neurons displayed low-threshold spikes (LTSs) and burst firing patterns. In 50% of cells tested, AVP induced a slowly rising, prolonged membrane depolarization and tonic firing, returning to burst firing upon recovery. AVP modulated hyperpolarization-activated LTSs by decreasing the time to the initial sodium spike at the onset of LTS, also increasing the duration of the afterdepolarization. Responses were blockable with a V(1a) receptor antagonist (Manning compound). Under voltage clamp, AVP induced a TTX-resistant, slowly rising, and prolonged (approximately 15 min) inward current (<40 pA). Current-voltage relationship (I-V) analyses of the AVP responses revealed a decrease in membrane conductance to 73.1 +/- 6.2% of control, with net AVP current reversing at -106 +/- 4 mV, and decreased inward rectification at negative potentials. These observations are consistent with an AVP-induced closure of an inwardly rectifying potassium conductance. On the basis of these in vitro observations, we suggest that the SCN vasopressinergic innervation of PVT is excitatory in nature, possibly releasing AVP with circadian rhythmicity and contributing to state-dependent firing patterns in PVT neurons over the sleep-wake cycle.