Role of L-Type Voltage-Gated Calcium Channels in Epileptiform Activity of Neurons

Epileptic discharges manifest in individual neurons as abnormal membrane potential fluctuations called paroxysmal depolarization shift (PDS). PDSs can combine into clusters that are accompanied by synchronous oscillations of the intracellular Ca2+ concentration ([Ca2+]i) in neurons. Here, we investigate the contribution of L-type voltage-gated calcium channels (VGCC) to epileptiform activity induced in cultured hippocampal neurons by GABA(A)R antagonist, bicuculline. Using KCl-induced depolarization, we determined the optimal effective doses of the blockers. Dihydropyridines (nifedipine and isradipine) at concentrations ≤ 10 μM demonstrate greater selectivity than the blockers from other groups (phenylalkylamines and benzothiazepines). However, high doses of dihydropyridines evoke an irreversible increase in [Ca2+]i in neurons and astrocytes. In turn, verapamil and diltiazem selectively block L-type VGCC in the range of 1–10 μM, whereas high doses of these drugs block other types of VGCC. We show that L-type VGCC blockade decreases the half-width and amplitude of bicuculline-induced [Ca2+]i oscillations. We also observe a decrease in the number of PDSs in a cluster and cluster duration. However, the pattern of individual PDSs and the frequency of the cluster occurrence change insignificantly. Thus, our results demonstrate that L-type VGCC contributes to maintaining the required [Ca2+]i level during oscillations, which appears to determine the number of PDSs in the cluster.

GABA Receptor Inhibition and Severe Hypoxia Induce a Paroxysmal Depolarization Shift in Goldfish Neurons

Mammalian neurons undergo rapid excitotoxic cell death when deprived of oxygen; however, the common goldfish (Carassius auratus) has the unique ability of surviving in oxygen-free waters. This organism utilizes γ-amino butyric acid (GABA) signaling to suppress excitatory glutamatergic activity during anoxic periods. Although GABAA receptor antagonists are not detrimental to the cell, co-inhibition of GABAA and GABAB receptors is detrimental by abolishing anoxia-induced neuro-protective mechanisms. In this article we show that blocking anoxic GABAergic neurotransmission induces seizure-like activity (SLA) analogous to a paroxysmal depolarization shift (PDS), with an elevation in action potential (AP) threshold and threshold current. The observed PDS was attributed to an increase in excitatory post-synaptic potential (EPSP) currents that are normally attenuated with decreasing oxygen levels. Furthermore, for the first time, we show that in addition to PDS, some neurons undergo depolarization block and do not generate AP despite a supra threshold membrane potential. In conclusion, our results indicate that with anoxia and absence of GABA receptor activity, telencephalic neurons of Carassius auratus manifest a paroxysmal depolarization shift, a key feature of epileptic discharge.

The Paroxysmal Depolarization Shift: Reconsidering Its Role in Epilepsy, Epileptogenesis and Beyond

Paroxysmal depolarization shifts (PDS) have been described by epileptologists for the first time several decades ago, but controversy still exists to date regarding their role in epilepsy. In addition to the initial view of a lack of such a role, seemingly opposing hypotheses on epileptogenic and anti-ictogenic effects of PDS have emerged. Hence, PDS may provide novel targets for epilepsy therapy. Evidence for the roles of PDS has often been obtained from investigations of the multi-unit correlate of PDS, an electrographic spike termed “interictal” because of its occurrence during seizure-free periods of epilepsy patients. Meanwhile, interictal spikes have been found to be associated with neuronal diseases other than epilepsy, e.g., Alzheimer’s disease, which may indicate a broader implication of PDS in neuropathologies. In this article, we give an introduction to PDS and review evidence that links PDS to pro- as well as anti-epileptic mechanisms, and to other types of neuronal dysfunction. The perturbation of neuronal membrane voltage and of intracellular Ca2+ that comes with PDS offers many conceivable pathomechanisms of neuronal dysfunction. Out of these, the operation of L-type voltage-gated calcium channels, which play a major role in coupling excitation to long-lasting neuronal changes, is addressed in detail.

Modulation of nickel-induced bursting with 4-aminopyridine in leech retzius nerve cells

Paroxysmal depolarization shift has been identified as a characteristic feature of the cellular basis of epilepsy. On Na+-dependent bursting, 1 mmol/l 4-aminopyridine (4-AP) produced a two-phase effect - a significant depolarization accompanied by an increase in the frequency of bursting, followed by repolarization along with a diminished frequency of bursting. Neither 1 ?mol/l apamin nor 150 nmol/l charybdotoxin (ChTX) elicited any significant effect on either bursting or standard conditions. Our results suggest that 4-AP affects the bursting indirectly by altering the excitability of the cell. The lack of effects of apamin and ChTX is probably due to channel insensitivity to these blockers in leech.

Activation of a Calcium-Activated Cation Current During Epileptiform Discharges and Its Possible Role in Sustaining Seizure-Like Events in Neocortical Slices

Epileptic seizures are composed of recurrent bursts of intense firing separated by periods of electrical quiescence. The mechanisms responsible for sustaining seizures and generating recurrent bursts are yet unclear. Using whole cell voltage recordings combined with intracellular calcium fluorescence imaging from bicuculline (BCC)-treated neocortical brain slices, I showed isolated paroxysmal depolarization shift (PDS) discharges were followed by a sustained afterdepolarization waveform (SADW) with an average peak amplitude of 3.3 ± 0.9 mV and average half-width of 6.2 ± 0.6 s. The SADW was mediated by the calcium-activated nonspecific cation current ( Ican) as it had a reversal potential of –33.1 ± 6.8 mV, was unaffected by changing the intracellular chloride concentrations, was markedly diminished by buffering [Ca2+]i with intracellular bis-( o-aminophenoxy)- N,N,N′,N′-tetraacetic acid (BAPTA), and was reversibly abolished by the Ican blocker flufenamic acid (FFA). The Ca2+ influx responsible for activation of Ican was mediated by both N-methyl-d-aspartate-receptor channels, voltage-gated calcium channels and, to a lesser extent, internal calcium stores. In addition to isolated PDS discharges, BCC-treated brain slices also produced seizure-like events, which were accompanied by a prolonged depolarizing waveform underlying individual ictal bursts. The similarities between the initial part of this waveform and the SADW and the fact it was markedly reduced by buffering [Ca2+]i with BAPTA strongly suggested it was mediated, at least in part, by Ican. Addition of FFA reversibly eliminated recurrent bursting, and transformed seizure-like events into isolated PDS responses. These results indicated Ican was activated during epileptiform discharges and probably participated in sustaining seizure-like events.

Abnormal neuronal responses during evolution of a penicillin epileptic focus in cat visual cortex

After defining the receptive fields of single units in cortical area 17 of anesthetized cats, recurrent on-off stimulation with bars of light of optimal configuration win from a second micropipette; Progressively, three distinct alterations of neuronal activity developed. The most longlasting and usually the earliest abnormality was an increase in the number and frequency of spikes comprising a neuron's response to stimuli that were effective prior to iontophoresis. This enhanced physiologic response (EPR) could be elicited from a cell independently of the discharge activity of an induced focus, but only with stimuli appropriate for the cell's receptive field. With additional iontophoresis an entirely new response developed, which was consistent with an extracellular paroxysmal depolarization shift (PDS). This high-frequency burst of spikes appeared only in association with an ECoG interictal potential. It could be triggered, however, by stimuli which were previously effective or ineffective, as well as occur spontaneously. Characteristics which further distinguished the PDS from EPR included a longer and more-variable latency, a longer recovery period, and a different sensitivity to changes of stimulus intensity. A period of response inhibition also accompanied each interictal potential and persisted with a variable duration afterward. It was most noticeable as an interruption in the activity of tonically responding neurons and was often present before the cell began to generate PDSs. It was concluded that the EPR represents a direct effect of penicillin on the cell or its immediate synaptic connections, while the PDS appears dependent on the altered interactions within a population of such affected cells. The inhibitory phenomenon, in addition, seems a result of projected influences from cells more fully involved with the developing focus. A dynamic model of the EPR-PDS relationship is proposed.