Tetrodotoxin induced calcium spikes: in vitro and in vivo studies of normal and deafferented Purkinje cells

Summary Tetrodotoxin (TTX) is widely used to block the sodium dependent action potential in excitable cells to study their other ionic properties. TTX applied outside, selectively blocks voltage dependent sodium channels and is thought to have no other effects. We report here that TTX, applied to slices of rat cerebellum, suppressed sodium spikes of the Purkinje cells and induced firing in bursts of slower spikes. This activity was blocked by cobalt (2 mM) or cadmium (0.2 mM) in the medium as well as by hyperpolarizing currents showing that the slow spikes were due to voltage dependent calcium channels. The membrane potential was not significantly changed by TTX and the spikes during the bursts had the same threshold potentials and peak spike amplitudes as the voltage and Ca²⁺ dependent dendritic spikes evoked by injected current before adding TTX. This indicated that no marked changes in the membrane conductances were produced by the TTX. Unlike the burst firing induced by removing extracellular sodium, the TTX induced bursts were not followed by a large hyperpolarization. The same kind of results were obtained with extracellular recording in the in-vivo preparation with TTX applied topically or by pressure near the recording sites. TTX induced burst firing was not due to blocking afferent inhibitory input to the PC, since bicuculline (10^-6 M) applied without TTX, produced only increased firing of fast action potentials and no bursts. The bursts could be arrested within 1 to 2 min by intravenously administering 2 mg/kg sodium pentobarbital, the blockage lasted from 5 to 15 min. These effects of TTX were not due to a contaminant as TTX from two different suppliers produced the same effects. A possible mechanism based on a decrease of intracellular free sodium is discussed.