Marco Martina

Functional differences in Na+ channel gating between fast‐spiking interneurones and principal neurones of rat hippocampus

1 GABAergic interneurones differ from glutamatergic principal neurones in their ability to discharge high‐frequency trains of action potentials without adaptation. To examine whether Na+ channel gating contributed to these differences, Na+ currents were recorded in nucleated patches from interneurones (dentate gyrus basket cells, BCs) and principal neurones (CA1 pyramidal cells, PCs) of rat hippocampal slices. 2 The voltage dependence of Na+ channel activation in BCs and PCs was similar. The slope factors of the activation curves, fitted with Boltzmann functions raised to the third power, were 11.5 and 11.8 mV, and the mid‐point potentials were −25.1 and −23.9 mV, respectively. 3 Whereas the time course of Na+ channel activation (−30 to +40 mV) was similar, the deactivation kinetics (−100 to −40 mV) were faster in BCs than in PCs (tail current decay time constants, 0.13 and 0.20 ms, respectively, at −40 mV). 4 Na+ channels in BCs and PCs differed in the voltage dependence of inactivation. The slope factors of the steady‐state inactivation curves fitted with Boltzmann functions were 6.7 and 10.7 mV, and the mid‐point potentials were −58.3 and −62.9 mV, respectively. 5 The onset of Na+ channel inactivation at −55 mV was slower in BCs than in PCs; the inactivation time constants were 18.6 and 9.3 ms, respectively. At more positive potentials the differences in inactivation onset were smaller. 6 The time course of recovery of Na+ channels from inactivation induced by a 30 ms pulse was fast and mono‐exponential (τ= 2.0 ms at −120 mV) in BCs, whereas it was slower and bi‐exponential in PCs (τ1= 2.0 ms and τ2= 133 ms; amplitude contribution of the slow component, 15%). 7 We conclude that Na+ channels of BCs and PCs differ in gating properties that contribute to the characteristic action potential patterns of the two types of neurones.

Scapuloperoneal spinal muscular atrophy and CMT2C are allelic disorders caused by alterations in TRPV4

Scapuloperoneal spinal muscular atrophy (SPSMA) and hereditary motor and sensory neuropathy type IIC (HMSN IIC, also known as HMSN2C or Charcot-Marie-Tooth disease type 2C (CMT2C)) are phenotypically heterogeneous disorders involving topographically distinct nerves and muscles. We originally described a large New England family of French-Canadian origin with SPSMA and an American family of English and Scottish descent with CMT2C. We mapped SPSMA and CMT2C risk loci to 12q24.1–q24.31 with an overlapping region between the two diseases. Further analysis reduced the CMT2C risk locus to a 4-Mb region. Here we report that SPSMA and CMT2C are allelic disorders caused by mutations in the gene encoding the transient receptor potential cation channel, subfamily V, member 4 (TRPV4). Functional analysis revealed that increased calcium channel activity is a distinct property of both SPSMA- and CMT2C-causing mutant proteins. Our findings link mutations in TRPV4 to altered calcium homeostasis and peripheral neuropathies, implying a pathogenic mechanism and possible options for therapy for these disorders.

Abnormalities in Hippocampal Functioning with Persistent Pain

Chronic pain patients exhibit increased anxiety, depression, and deficits in learning and memory. Yet how persistent pain affects the key brain area regulating these behaviors, the hippocampus, has remained minimally explored. In this study we investigated the impact of spared nerve injury (SNI) neuropathic pain in mice on hippocampal-dependent behavior and underlying cellular and molecular changes. In parallel, we measured the hippocampal volume of three groups of chronic pain patients. We found that SNI animals were unable to extinguish contextual fear and showed increased anxiety-like behavior. Additionally, SNI mice compared with Sham animals exhibited hippocampal (1) reduced extracellular signal-regulated kinase expression and phosphorylation, (2) decreased neurogenesis, and (3) altered short-term synaptic plasticity. To relate the observed hippocampal abnormalities with human chronic pain, we measured the volume of human hippocampus in chronic back pain (CBP), complex regional pain syndrome (CRPS), and osteoarthritis patients (OA). Compared with controls, CBP and CRPS, but not OA, had significantly less bilateral hippocampal volume. These results indicate that hippocampus-mediated behavior, synaptic plasticity, and neurogenesis are abnormal in neuropathic rodents. The changes may be related to the reduction in hippocampal volume we see in chronic pain patients, and these abnormalities may underlie learning and emotional deficits commonly observed in such patients.

R-Type Calcium Channels Contribute to Afterdepolarization and Bursting in Hippocampal CA1 Pyramidal Neurons

Action potentials in pyramidal neurons are typically followed by an afterdepolarization (ADP), which in many cells contributes to intrinsic burst firing. Despite the ubiquity of this common excitable property, the responsible ion channels have not been identified. Using current-clamp recordings in hippocampal slices, we find that the ADP in CA1 pyramidal neurons is mediated by an Ni2+-sensitive calcium tail current. Voltage-clamp experiments indicate that the Ni2+-sensitive current has a pharmacological and biophysical profile consistent with R-type calcium channels. These channels are available at the resting potential, are activated by the action potential, and remain open long enough to drive the ADP. Because the ADP correlates directly with burst firing in CA1 neurons, R-type calcium channels are crucial to this important cellular behavior, which is known to encode hippocampal place fields and enhance synaptic plasticity.

Dendritic Mechanisms Underlying Rapid Synaptic Activation of Fast-Spiking Hippocampal Interneurons

Dendrites Shape Interneuron Firing Basket cells, a group of fast-spiking inhibitory interneurons, play an important part in the function of neuronal networks. The mechanisms underlying the high temporal precision and short latency of basket cell activity are unclear. Hu et al. (p. 52, published online 3 December) investigated dendrite functions in fast-spiking hippocampal basket cells and found that action potentials are initiated in the axon and propagate back into the dendrites without activity dependence but with strongly reduced amplitude. This is very different from what has been observed previously in widely investigated pyramidal cell dendrites, probably due to the high potassium to sodium conductance ratios in the dendrites of the interneurons. These dendritic mechanisms can explain the high-frequency firing and precise timing of basket cells seen in network activity in vivo. Potassium channel enrichment in the dendrites of hippocampal basket cells defines a mechanism of neural network function. Fast-spiking, parvalbumin-expressing basket cells (BCs) are important for feedforward and feedback inhibition. During network activity, BCs respond with short latency and high temporal precision. It is thought that the specific properties of input synapses are responsible for rapid recruitment. However, a potential contribution of active dendritic conductances has not been addressed. We combined confocal imaging and patch-clamp techniques to obtain simultaneous somatodendritic recordings from BCs. Action potentials were initiated in the BC axon and backpropagated into the dendrites with reduced amplitude and little activity dependence. These properties were explained by a high K+ to Na+ conductance ratio in BC dendrites. Computational analysis indicated that dendritic K+ channels convey unique integration properties to BCs, leading to the rapid and temporally precise activation by excitatory inputs.

Gating, modulation and subunit composition of voltage‐gated K+ channels in dendritic inhibitory interneurones of rat hippocampus

GABAergic interneurones are diverse in their morphological and functional properties. Perisomatic inhibitory cells show fast spiking during sustained current injection, whereas dendritic inhibitory cells fire action potentials with lower frequency. We examined functional and molecular properties of K+ channels in interneurones with horizontal dendrites in stratum oriens‐alveus (OA) of the hippocampal CA1 region, which mainly comprise somatostatin‐positive dendritic inhibitory cells. Voltage‐gated K+ currents in nucleated patches isolated from OA interneurones consisted of three major components: a fast delayed rectifier K+ current component that was highly sensitive to external 4‐aminopyridine (4‐AP) and tetraethylammonium (TEA) (half‐maximal inhibitory concentrations < 0.1 mm for both blockers), a slow delayed rectifier K+ current component that was sensitive to high concentrations of TEA, but insensitive to 4‐AP, and a rapidly inactivating A‐type K+ current component that was blocked by high concentrations of 4‐AP, but resistant to TEA. The relative contributions of these components to the macroscopic K+ current were estimated as 57 ± 5, 25 ± 6, and 19 ± 2 %, respectively. Dendrotoxin, a selective blocker of Kv1 channels had only minimal effects on K+ currents in nucleated patches. Coapplication of the membrane‐permeant cAMP analogue 8‐(4‐chlorophenylthio)‐adenosine 3′:5′‐cyclic monophosphate (cpt‐cAMP) and the phosphodiesterase blocker isobutyl‐methylxanthine (IBMX) resulted in a selective inhibition of the fast delayed rectifier K+ current component. This inhibition was absent in the presence of the protein kinase A (PKA) inhibitor H‐89, implying the involvement of PKA‐mediated phosphorylation. Single‐cell reverse transcription‐polymerase chain reaction (RT‐PCR) analysis revealed a high abundance of Kv3.2 mRNA in OA interneurones, whereas the expression level of Kv3.1 mRNA was markedly lower. Similarly, RT‐PCR analysis showed a high abundance of Kv4.3 mRNA, whereas Kv4.2 mRNA was undetectable. This suggests that the fast delayed rectifier K+ current and the A‐type K+ current component are mediated predominantly by homomeric Kv3.2 and Kv4.3 channels. Selective modulation of Kv3.2 channels in OA interneurones by cAMP is likely to be an important factor regulating the activity of dendritic inhibitory cells in principal neurone‐interneurone microcircuits.

The unipolar brush cell: a remarkable neuron finally receiving deserved attention.

Unipolar brush cells (UBC) are small, glutamatergic neurons residing in the granular layer of the cerebellar cortex and the granule cell domain of the cochlear nuclear complex. Recent studies indicate that this neuronal class consists of three or more subsets characterized by distinct chemical phenotypes, as well as by intrinsic properties that may shape their synaptic responses and firing patterns. Yet, all UBCs have a unique morphology, as both the dendritic brush and the large endings of the axonal branches participate in the formation of glomeruli. Although UBCs and granule cells may share the same excitatory and inhibitory inputs, the two cell types are distinctively differentiated. Typically, whereas the granule cell has 4-5 dendrites that are innervated by different mossy fibers, and an axon that divides only once to form parallel fibers after ascending to the molecular layer, the UBC has but one short dendrite whose brush engages in synaptic contact with a single mossy fiber terminal, and an axon that branches locally in the granular layer; branches of UBC axons form a non-canonical, cortex-intrinsic category of mossy fibers synapsing with granule cells and other UBCs. This is thought to generate a feed-forward amplification of single mossy fiber afferent signals that would reach the overlying Purkinje cells via ascending granule cell axons and their parallel fibers. In sharp contrast to other classes of cerebellar neurons, UBCs are not distributed homogeneously across cerebellar lobules, and subsets of UBCs also show different, albeit overlapping, distributions. UBCs are conspicuously rare in the expansive lateral cerebellar areas targeted by the cortico-ponto-cerebellar pathway, while they are a constant component of the vermis and the flocculonodular lobe. The presence of UBCs in cerebellar regions involved in the sensorimotor processes that regulate body, head and eye position, as well as in regions of the cochlear nucleus that process sensorimotor information suggests a key role in these critical functions; it also invites further efforts to clarify the cellular biology of the UBCs and their specific functions in the neuronal microcircuits in which they are embedded. High density of UBCs in specific regions of the cerebellar cortex is a feature largely conserved across mammals and suggests an involvement of these neurons in fundamental aspects of the input/output organization as well as in clinical manifestation of focal cerebellar disease.

Chronic neuropathic pain-like behavior correlates with IL-1β expression and disrupts cytokine interactions in the hippocampus.

Summary IL‐1β expression in the contralateral hippocampus coincides with neuropathic pain behavior in rats, and the correlations between hippocampal IL‐1β and IL‐1ra or IL‐6 are lost. ABSTRACT We have proposed that neuropathic pain engages emotional learning, suggesting the involvement of the hippocampus. Because cytokines in the periphery contribute to induction and maintenance of neuropathic pain but might also participate centrally, we used 2 neuropathic pain models, chronic constriction injury (CCI) and spared nerve injury (SNI), to investigate the temporal profile of hippocampal cytokine gene expression in 2 rat strains that show different postinjury behavioral threshold sensitivities. SNI induced long‐lasting allodynia in both strains, while CCI induced allodynia with time‐dependent recovery in Sprague Dawley (SD) and no allodynia in Wistar Kyoto (WK) rats. In WK rats, only SNI induced sustained upregulation of hippocampal interleukin (IL)‐1β, while IL‐6 expression was transiently increased and no significant changes in IL‐1ra expression were detected. Conversely, in SD rats, SNI resulted in sustained and robust increased hippocampal IL‐1β expression, which was only transient in rats with CCI. In this strain, IL‐6 expression was not affected in any of the 2 injury models and IL‐1ra expression was significantly increased in rats with SNI or CCI at late phases. We found that the degree and development of neuropathic pain depend on the specific nerve injury model and rat strain; that hippocampal IL‐1β mRNA levels correlate with neuropathic pain behavior; that, in contrast to sham‐operated animals, there are no correlations between hippocampal IL‐1β and IL‐1ra or IL‐6 in neuropathic rats; and that alterations in cytokine expression are restricted to the hippocampus contralateral to the injury side, again implying that the observed changes reflect nociception.

Sodium Currents Activate without a Hodgkin and Huxley-Type Delay in Central Mammalian Neurons

Hodgkin and Huxley established that sodium currents in the squid giant axons activate after a delay, which is explained by the model of a channel with three identical independent gates that all have to open before the channel can pass current (the HH model). It is assumed that this model can adequately describe the sodium current activation time course in all mammalian central neurons, although there is no experimental evidence to support such a conjecture. We performed high temporal resolution studies of sodium currents gating in three types of central neurons. The results show that, within the tested voltage range from -55 to -35 mV, in all of these neurons, the activation time course of the current could be fit, after a brief delay, with a monoexponential function. The duration of delay from the start of the voltage command to the start of the extrapolated monoexponential fit was much smaller than predicted by the HH model. For example, in prefrontal cortex pyramidal neurons, at -46 mV and 12°C, the observed average delay was 140 μs versus the 740 μs predicted by the two-gate HH model and the 1180 μs predicted by the three-gate HH model. These results can be explained by a model with two closed states and one open state. In this model, the transition between two closed states is approximately five times faster than the transition between the second closed state and the open state. This model captures all major properties of the sodium current activation. In addition, the proposed model reproduces the observed action potential shape more accurately than the traditional HH model.

Differential Expression of TASK Channels between Horizontal Interneurons and Pyramidal Cells of Rat Hippocampus

Among the electrophysiological properties differentiating stratum oriens horizontal interneurons from pyramidal neurons of the CA1 hippocampal subfield are the more depolarized resting potential and the higher input resistance; additionally, these interneurons are also less sensitive to ischemic damage than pyramidal cells. A differential expression of pH-sensitive leakage potassium channels (TASK) could contribute to all of these differences. To test this hypothesis, we studied the expression and properties of TASK channels in the two cell types. Electrophysiological recordings from acute slices showed that barium- and bupivacaine-sensitive TASK currents were detectable in pyramidal cells but not in interneurons and that extracellular acidification caused a much stronger depolarization in pyramidal cells than in interneurons. This pyramidal cell depolarization was paralleled by an increase of the input resistance, suggesting the blockade of a background conductance. Single-cell reverse transcription-PCR experiments showed that the expression profile of TASK channels differ between the two cell types and suggested that these channels mediate an important share of the leakage current of pyramidal cells. We suggest that the different expression of TASK channels in these cell types contribute to their electrophysiological differences and may result in cell-specific sensitivity to extracellular acidification in conditions such as epilepsy and ischemia.