Biomedical subjects

Fernando R Fernandez

Publications and source records attributed to Fernando R Fernandez.

7 recordsLinked to original sources

PubMed2006

Importance of K+-dependent Na+/Ca2+-exchanger 2, NCKX2, in motor learning and memory.

Plasma membrane Na+/Ca2+-exchangers play a predominant role in Ca2+ extrusion in brain. Neurons express several different Na+/Ca2+-exchangers belonging to both the K+-independent NCX family and the K+-dependent NCKX family. The unique contributions of each of these proteins to neuronal Ca2+ homeostasis and/or physiology remain largely unexplored. To address this question, we generated mice in which the gene encoding the abundant neuronal K+ -dependent Na+/Ca2+-exchanger protein, NCKX2, was knocked out. Analysis of these animals revealed a significant reduction in Ca2+ flux in cortical neurons, a profound loss of long term potentiation and an increase in long term depression at hippocampal Schaffer/CA1 synapses, and clear deficits in specific tests of motor learning and spatial working memory. Surprisingly, there was no obvious loss of photoreceptor function in cones, where expression of the NCKX2 protein had been reported previously. These data emphasize the critical and non-redundant role of NCKX2 in the local control of neuronal [Ca2+] that is essential for the development of synaptic plasticity associated with learning and memory.

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PubMed2005

A C-terminal domain directs Kv3.3 channels to dendrites.

Pyramidal neurons of the electrosensory lateral line lobe (ELL) of Apteronotus leptorhynchus express Kv3-type voltage-gated potassium channels that give rise to high-threshold currents at the somatic and dendritic levels. Two members of the Kv3 channel family, AptKv3.1 and AptKv3.3, are coexpressed in these neurons. AptKv3.3 channels are expressed at uniformly high levels in each of four ELL segments, whereas AptKv3.1 channels appear to be expressed in a graded manner with higher levels of expression in segments that process high-frequency electrosensory signals. Immunohistochemical and recombinant channel expression studies show a differential distribution of these two channels in the dendrites of ELL pyramidal neurons. AptKv3.1 is concentrated in somas and proximal dendrites, whereas AptKv3.3 is distributed throughout the full extent of the large dendritic tree. Recombinant channel expression of AptKv3 channels through in vivo viral injections allowed directed retargeting of AptKv3 subtypes over the somadendritic axis, revealing that the sequence responsible for targeting channels to distal dendrites lies within the C-terminal domain of the AptKv3.3 protein. The targeting domain includes a consensus sequence predicted to bind to a PDZ (postsynaptic density-95/Discs large/zona occludens-1)-type protein-protein interaction motif. These findings reveal that different functional roles for Kv3 potassium channels at the somatic and dendritic level of a sensory neuron are attained through specific targeting that selectively distributes Kv3.3 channels to the dendritic compartment.

Amino Acid Sequence↗

PubMed2005

A-type and T-type currents interact to produce a novel spike latency-voltage relationship in cerebellar stellate cells.

The modification of first-spike latencies by low-threshold and inactivating K+ currents (IA) have important implications in neuronal coding and synaptic integration. To date, cells in which first-spike latency characteristics have been analyzed have shown that increased hyperpolarization results in longer first-spike latencies, producing a monotonic relationship between first-spike latency and membrane voltage. Previous work has established that cerebellar stellate cells express members of the Kv4 potassium channel subfamily, which underlie IA in many central neurons. Spike timing in stellate cells could be particularly important to cerebellar output, because the discharge of even single spikes can significantly delay spike discharge in postsynaptic Purkinje cells. In the present work, we studied the first-spike latency characteristics of stellate cells. We show that first-spike latency is nonmonotonic, such that intermediate levels of prehyperpolarization produce the longest spike latencies, whereas greater hyperpolarization or depolarization reduces spike latency. Moreover, the range of first-spike latency values can be substantial in spanning 20-128 ms with preceding membrane shifts of <10 mV. Using patch clamp and modeling, we illustrate that spike latency characteristics are the product of an interplay between IA and low-threshold calcium current (IT) that requires a steady-state difference in the inactivation parameters of the currents. Furthermore, we show that the unique first-spike latency characteristics of stellate cells have important implications for the integration of coincident IPSPs and EPSPs, such that inhibition can shift first-spike latency to differentially modulate the probability of firing.

Action Potentials↗

PubMed2005

Dendritic Na+ current inactivation can increase cell excitability by delaying a somatic depolarizing afterpotential.

Many central neurons support active dendritic spike backpropagation mediated by voltage-gated currents. Active spikes in dendrites have been shown capable of providing feedback to the soma to influence somatic excitability and firing dynamics through a depolarizing afterpotential (DAP). In pyramidal cells of the electrosensory lobe of weakly electric fish, Na(+) spikes in dendrites undergo a frequency-dependent broadening that enhances the DAP to increase somatic firing frequency. We use a combination of dynamical analysis and electrophysiological recordings to demonstrate that spike broadening in dendrites is primarily caused by a cumulative inactivation of dendritic Na(+) current. We further show that a reduction in dendritic Na(+) current increases excitability by decreasing the interspike interval and promoting burst firing. This process arises when inactivation of dendritic Na(+) current shifts the latency of the dendritic spike to delay the arrival of the DAP sufficiently to increase its impact on somatic membrane potential despite a reduction in dendritic excitability. Furthermore, the relationship between dendritic Na(+) current density and somatic excitability is nonmonotonic, as intermediate levels of dendritic Na(+) current exert the greatest excitatory influence. These results reveal that temporal shifts in dendritic spike firing provide a novel means for backpropagating spikes to influence the final output of a cell.

Action Potentials↗

PubMed2005

High-threshold K+ current increases gain by offsetting a frequency-dependent increase in low-threshold K+ current.

High-frequency firing neurons are found in numerous central systems, including the auditory brainstem, thalamus, hippocampus, and neocortex. The kinetics of high-threshold K+ currents (IK(HT)) from the Kv3 subfamily has led to the proposal that these channels offset cumulative Na+ current inactivation and stabilize tonic high-frequency firing. However, all high-frequency firing neurons, examined to date, also express low-threshold K+ currents (IK(LT)) that have slower kinetics and play an important role in setting the subthreshold and filtering properties of the neuron. IK(LT) has also been shown to dampen excitability and is therefore likely to oppose high-frequency firing. In this study, we examined the role of IK(HT) in pyramidal cells of the electrosensory lobe of weakly electric fish, which are characterized by high-frequency firing, a very wide frequency range, and high levels of IK(HT). In particular, we examined the mechanisms that allow IK(HT) to set the gain of the F-I relationship by interacting with another low-threshold K+ current. We found that IK(HT) increases the gain of the F-I relationship and influences spike waveform almost exclusively in the high-frequency firing range. The frequency dependence arises from IK(HT) influencing both the IK(LT) and Na+ currents. IK(HT) thus plays a significant role in stabilizing high-frequency firing by preventing a steady-state accumulation of IK(LT) that is as important as preventing Na+ current inactivation.

Action Potentials↗

PubMed2004

Releasing the peri-neuronal net to patch-clamp neurons in adult CNS.

The extracellular matrix of adult neural tissue contains chondroitin sulphated proteogylcans that form a dense peri-neuronal net surrounding the cell body and proximal dendrites of many neuronal classes. Development of the peri-neuronal net beyond approximately postnatal day 17 obscures visualization and often access by patch electrodes to neuronal membranes with the result that patch clamp recordings are most readily obtained from early postnatal animals. We describe a technique in which the surface tension of a sucrose-based medium promotes partial dissociation of thin tissue slices from adult tissue. Surface tension spreads the tissue and loosens the peri-neuronal net from neuronal membranes within minutes and in the absence of proteolytic enzymes. Furthermore, the extent of dissociation can be controlled so as to maintain the overall slice structure and allow identification of specific cell classes. Excellent structural preservation of neurons and dendrites can be obtained and full access by patch electrodes made possible for current- or voltage-clamp recordings in tissue well beyond the development of peri-neuronal nets. We demonstrate the feasibility of using this approach through patch recordings from neurons in the brainstem and cerebellum of adult gymnotiform fish and in deep cerebellar nuclei of rats as old as 6 months.

Animals↗

PubMed2003

Inactivation of Kv3.3 potassium channels in heterologous expression systems.

Kv3.3 K+ channels are believed to incorporate an NH2-terminal domain to produce an intermediate rate of inactivation relative to the fast inactivating K+ channels Kv3.4 and Kv1.4. The rate of Kv3.3 inactivation has, however, been difficult to establish given problems in obtaining consistent rates of inactivation in expression systems. This study characterized the properties of AptKv3.3, the teleost homologue of Kv3.3, when expressed in Chinese hamster ovary (CHO) or human embryonic kidney (HEK) cells. We show that the properties of AptKv3.3 differ significantly between CHO and HEK cells, with the largest difference occurring in the rate and voltage dependence of inactivation. While AptKv3.3 in CHO cells showed a fast and voltage-dependent rate of inactivation consistent with N-type inactivation, currents in HEK cells showed rates of inactivation that were voltage-independent and more consistent with a slower C-type inactivation. Examination of the mRNA sequence revealed that the first methionine start site had a weak Kozak consensus sequence, suggesting that the lack of inactivation in HEK cells could be due to translation at a second methionine start site downstream of the NH2-terminal coding region. Mutating the nucleotide sequence surrounding the first methionine start site to one more closely resembling a Kozak consensus sequence produced currents that inactivated with a fast and voltage-dependent rate of inactivation in both CHO and HEK cells. These results indicate that under the appropriate conditions Kv3.3 channels can exhibit fast and reliable inactivation that approaches that more typically expected of "A"-type K+ currents.

Amino Acid Sequence↗