Study of the effects of ethanol on the experimental model of Nickel-induced epileptiform activity in retzius neurons of the leech haemopis sanguisuga.

Thesis Report.

ABSTRACT: Epileptic seizures are based on the appearance of paroxysmal depolarization shifts (PDS) which are synchronized in many neurons. Mechanisms underlying PDS and seizures are still not understood. Essential mechanisms of epileptic activity in nervous systems are thought to be identical in whatever part of the human or animal nervous system epileptic activity appears. Invertebrate systems have proved to be quite useful for the development of an understanding of some processes in the central nervous system. Because invertebrate neurons have been so accessible to experimentation, it has been possible to explore in detail the basic mechanisms controlling neuronal excitability using these cells and to make some useful predictions about electrophysiological mechanisms that may be present in central neurons. As in other invertebrate preparations, it has been possible to investigate the cellular basis for epileptiform activity in Retzius nerve cells comprising the relevant neuronal circuits and to determine their cellular and synaptic properties. In the present study we have examined the effects of ethanol on Ni2+-induced bursting of leech Retzius nerve cells. Saline with 3 mmol/l NiCl2 induced spontaneous bursting activity, characterized by rapid depolarizations to a plateau level during which bursts of action potentials occurred. To test for the mechanism of bursting initiation external Na+ was completely removed. Removal of external Na+ in presence of 3 mmol/l NiCl2 terminated the bursting activity. Application of 2 % ethanol solution significantly decreased the bursting frequency, duration and amplitude of depolarization plateaus, and the number of spikes per plateau. The suppression of bursting was in the order of 2.0 %> 1.0 %> 0.3 %>0.1 %. Clearly, two types of bursting viz. synaptically triggered- Ca2+-dependent PDSs and nonsyptically triggered- Na+-dependent PDSs were first researched. Although two major types of PDSs are known to exist, special focus on Ca2+-dependent PDSs has overshadowed the Na+-dependent PDSs. The Retzius nerve cell of the leech offers the possibility to study the Na+-dependent PDS-like events which occur during the block of calcium channels by 3 mmol/l Ni2+, as well as a possible role of altered potassium channel dynamics in initiation of these PDSs. In this regard, we have checked the effects of K+ channel blockers 4-Aminopyridine (4-AP), apamin, and charybdotoxin (ChTX) on nickel induced bursting of the Retzius nerve cells using classical electrophysiological recordings and current clamp measurements. 1 mmol/l 4-AP produced a two-phase effect - a significant depolarization accompanied by an increase in frequency of bursting, followed by repolarization along with diminished frequency of bursting. Similar effect of 4-AP was recorded in standard conditions. 1 μmol/l apamin and 150 nmol/l charybdotoxin (ChTX) did not elicit any significant effect on either bursting or in standard conditions. Our results suggest that 4-AP potentiated the bursting indirectly by altering excitability of the cell. Lack of effects of apamin and ChTX is probably due to channel insensitiveness to these blockers in leech. Increase of input membrane resistance in Ni Ri suggest the block of voltage gated Ca2+-channels or Ca2+ activated K+ channels or both, and the decrease during peak of the plateau indicates opening of Na+ channels. Finally, current clamp study shows increase in input membrane resistance in both case of applied ethanol in standard physiological saline and Ni2+ induced bursting; corroborating the idea that ethanol might have suppressed the bursting by acting via Na+ channels. Key words: Paroxysmal depolarization shift, bursting, ethanol, 4-AP, charybdotoxin, apamin, input membrane resistance, Retzius nerve cells, leech.

Eng.