Protection from noise-induced hearing loss by Kv2.2 potassium currents in the central medial olivocochlear system.

The central auditory brainstem provides an efferent projection known as the medial olivocochlear (MOC) system, which regulates the cochlear amplifier and mediates protection on exposure to loud sound. It arises from neurons of the ventral nucleus of the trapezoid body (VNTB), so control of neuronal excitability in this pathway has profound effects on hearing. The VNTB and the medial nucleus of the trapezoid body are the only sites of expression for the Kv2.2 voltage-gated potassium channel in the auditory brainstem, consistent with a specialized function of these channels. In the absence of unambiguous antagonists, we used recombinant and transgenic methods to examine how Kv2.2 contributes to MOC efferent function. Viral gene transfer of dominant-negative Kv2.2 in wild-type mice suppressed outward K(+) currents, increasing action potential (AP) half-width and reducing repetitive firing. Similarly, VNTB neurons from Kv2.2 knock-out mice (Kv2.2KO) also showed increased AP duration. Control experiments established that Kv2.2 was not expressed in the cochlea, so any changes in auditory function in the Kv2.2KO mouse must be of central origin. Further, in vivo recordings of auditory brainstem responses revealed that these Kv2.2KO mice were more susceptible to noise-induced hearing loss. We conclude that Kv2.2 regulates neuronal excitability in these brainstem nuclei by maintaining short APs and enhancing high-frequency firing. This safeguards efferent MOC firing during high-intensity sounds and is crucial in the mediation of protection after auditory overexposure.

Early interneuron dysfunction in ALS: insights from a mutant sod1 zebrafish model.

OBJECTIVE: To determine, when, how, and which neurons initiate the onset of pathophysiology in amyotrophic lateral sclerosis (ALS) using a transgenic mutant sod1 zebrafish model and identify neuroprotective drugs. METHODS: Proteinopathies such as ALS involve mutant proteins that misfold and activate the heat shock stress response (HSR). The HSR is indicative of neuronal stress, and we used a fluorescent hsp70-DsRed reporter in our transgenic zebrafish to track neuronal stress and to measure functional changes in neurons and muscle over the course of the disease. RESULTS: We show that mutant sod1 fish first exhibited the HSR in glycinergic interneurons at 24 hours postfertilization (hpf). By 96 hpf, we observed a significant reduction in spontaneous glycinergic currents induced in spinal motor neurons. The loss of inhibition was followed by increased stress in the motor neurons of symptomatic adults and concurrent morphological changes at the neuromuscular junction (NMJ) indicative of denervation. Riluzole, the only approved ALS drug and apomorphine, an NRF2 activator, reduced the observed early neuronal stress response. INTERPRETATION: The earliest event in the pathophysiology of ALS in the mutant sod1 zebrafish model involves neuronal stress in inhibitory interneurons, resulting from mutant Sod1 expression. This is followed by a reduction in inhibitory input to motor neurons. The loss of inhibitory input may contribute to the later development of neuronal stress in motor neurons and concurrent inability to maintain the NMJ. Riluzole, the approved drug for use in ALS, modulates neuronal stress in interneurons, indicating a novel mechanism of riluzole action.

Pacemaker and plateau potentials shape output of a developing locomotor network.

BACKGROUND: During development, spinal networks undergo an intense period of maturation in which immature forms of motor behavior are observed. Such behaviors are transient, giving way to more mature activity as development proceeds. The processes governing age-specific transitions in motor behavior are not fully understood. RESULTS: Using in vivo patch clamp electrophysiology, we have characterized ionic conductances and firing patterns of developing zebrafish spinal neurons. We find that a kernel of spinal interneurons, the ipsilateral caudal (IC) cells, generate inherent bursting activity that depends upon a persistent sodium current (I(NaP)). We further show that developmental transitions in motor behavior are accompanied by changes in IC cell bursting: during early life, these cells generate low frequency membrane oscillations that likely drive "coiling," an immature form of motor output. As fish mature to swimming stages, IC cells switch to a sustained mode of bursting that permits generation of high-frequency oscillations during locomotion. Finally, we find that perturbation of IC cell bursting disrupts motor output at both coiling and swimming stages. CONCLUSIONS: Our results suggest that neurons with unique bursting characteristics are a fundamental component of developing motor networks. During development, these may shape network output and promote stage-specific reconfigurations in motor behavior.

Nitric oxide is an activity-dependent regulator of target neuron intrinsic excitability.

Activity-dependent changes in synaptic strength are well established as mediating long-term plasticity underlying learning and memory, but modulation of target neuron excitability could complement changes in synaptic strength and regulate network activity. It is thought that homeostatic mechanisms match intrinsic excitability to the incoming synaptic drive, but evidence for involvement of voltage-gated conductances is sparse. Here, we show that glutamatergic synaptic activity modulates target neuron excitability and switches the basis of action potential repolarization from Kv3 to Kv2 potassium channel dominance, thereby adjusting neuronal signaling between low and high activity states, respectively. This nitric oxide-mediated signaling dramatically increases Kv2 currents in both the auditory brain stem and hippocampus (>3-fold) transforming synaptic integration and information transmission but with only modest changes in action potential waveform. We conclude that nitric oxide is a homeostatic regulator, tuning neuronal excitability to the recent history of excitatory synaptic inputs over intervals of minutes to hours.

Regulation of Kv channel expression and neuronal excitability in rat medial nucleus of the trapezoid body maintained in organotypic culture.

Principal neurons of the medial nucleus of the trapezoid body (MNTB) express a spectrum of voltage-dependent K(+) conductances mediated by Kv1-Kv4 channels, which shape action potential (AP) firing and regulate intrinsic excitability. Postsynaptic factors influencing expression of Kv channels were explored using organotypic cultures of brainstem prepared from P9-P12 rats and maintained in either low (5 mm, low-K) or high (25 mm, high-K) [K(+)](o) medium. Whole cell patch-clamp recordings were made after 7-28 days in vitro. MNTB neurons cultured in high-K medium maintained a single AP firing phenotype, while low-K cultures had smaller K(+) currents, enhanced excitability and fired multiple APs. The calyx of Held inputs degenerated within 3 days in culture, having lost their major afferent input; this preparation of calyx-free MNTB neurons allowed the effects of postsynaptic depolarisation to be studied with minimal synaptic activity. The depolarization caused by the high-K aCSF only transiently increased spontaneous AP firing (<2 min) and did not measurably increase synaptic activity. Chronic depolarization in high-K cultures raised basal levels of [Ca(2+)](i), increased Kv3 currents and shortened AP half-widths. These events relied on raised [Ca(2+)](i), mediated by influx through voltage-gated calcium channels (VGCCs) and release from intracellular stores, causing an increase in cAMP-response element binding protein (CREB) phosphorylation. Block of VGCCs or of CREB function suppressed Kv3 currents, increased AP duration, and reduced Kv3.3 and c-fos expression. Real-time PCR revealed higher Kv3.3 and Kv1.1 mRNA in high-K compared to low-K cultures, although the increased Kv1.1 mRNA was mediated by a CREB-independent mechanism. We conclude that Kv channel expression and hence the intrinsic membrane properties of MNTB neurons are homeostatically regulated by [Ca(2+)](i)-dependent mechanisms and influenced by sustained depolarization of the resting membrane potential.

Nitric oxide is a volume transmitter regulating postsynaptic excitability at a glutamatergic synapse.

Neuronal nitric oxide synthase (nNOS) is broadly expressed in the brain and associated with synaptic plasticity through NMDAR-mediated calcium influx. However, its physiological activation and the mechanisms by which nitric oxide (NO) influences synaptic transmission have proved elusive. Here, we exploit the unique input-specificity of the calyx of Held to characterize NO modulation at this glutamatergic synapse in the auditory pathway. NO is generated in an activity-dependent manner by MNTB principal neurons receiving a calyceal synaptic input. It acts in the target neuron and adjacent inactive neurons to modulate excitability and synaptic efficacy, inhibiting postsynaptic Kv3 potassium currents (via phosphorylation), reducing EPSCs and so increasing action potential duration and reducing transmission fidelity. We conclude that NO serves as a volume transmitter and slow dynamic modulator, integrating spontaneous and evoked neuronal firing, thereby providing an index of global activity and regulating information transmission across a population of active and inactive neurons.

Dopamine D1 receptor inhibition of NMDA receptor currents mediated by tyrosine kinase-dependent receptor trafficking in neonatal rat striatum.

NMDA receptors are of particular importance in the control of synaptic strength and integration of synaptic activity. Dopamine receptor modulation of NMDA receptors in neonatal striatum may influence the efficacy of synaptic transmission in the cortico-striatal pathway and if so, this modulation will affect the behaviour of the basal ganglia network. Here, we show that in acute brain slices of neonatal (P7) rat striatum the dopamine D1 receptor agonist SKF-82958 significantly decreases NMDA receptor currents in patch-clamp whole-cell recordings. This inhibition is not abolished by application of a G protein inhibitor (GDP-beta-S) or irreversible G protein activator (GTP-gamma-S) suggesting a G protein-independent mechanism. In addition, intracellular application of protein tyrosine kinase inhibitors (lavendustin A or PP2) abolished D1 inhibition of NMDA currents. In contrast, in older animals (P28) D1 receptor activation produces a potentiation of the NMDA response which suggests there is a developmental switch in D1 modulation of striatal NMDA receptors. Single-channel recordings show that direct D1 receptor inhibition of NMDA receptors cannot be observed in isolated membrane patches. We hypothesize that D1 inhibition in whole-cell recordings from neonatal rats may be mediated by a change in NMDA receptor trafficking. Consistent with this hypothesis, intracellular application of a dynamin inhibitory peptide (QVPSRPNRAP) abolished D1 inhibition of NMDA receptor currents. We therefore conclude that a tyrosine kinase-dependent alteration of NMDA receptor trafficking underlies D1 dopamine receptor-mediated down-regulation of NMDA receptor currents in medium spiny neurons of neonatal rat striatum.