Abstracts

Rationale: Leptin inhibits the Ca²⁺ oscillations and epileptiform-like activity observed in hippocampal neurons when exposed to a Mg²⁺-free extracellular solution. To further test leptin’s potential as an anticonvulsant, we examined the effects of leptin on neuronal bursting induced by a Mg²⁺-free extracellular solution. We also examined the effects of leptin on action potentials, input resistance, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) currents, and N-methyl-D-aspartate (NMDA) currents.Methods: Current and voltage clamp recordings were obtained from cultured embryonic mouse hippocampal neurons using the whole-cell patch clamp technique. The effects of leptin on action potentials, input resistance, and AMPA currents were examined with 1 mM extracellular Mg²⁺, but the effect on NMDA currents was examined in a Mg²⁺-free extracellular solution containing 10 μM glycine. Neurons were subjected to 700 ms current steps ranging from -40 pA to +40 pA in 10 pA increments to generate action potentials and to determine input resistance. AMPA and NMDA were applied using a flow-tube system. Results: Cultured hippocampal neurons generally fired isolated action potentials with 1 mM Mg²⁺ in the extracellular solution. When exposed to a Mg²⁺-free extracellular solution, neurons produced bursts of action potentials superimposed on 0.5-2 s depolarizing envelopes resembling paroxysmal depolarizing shifts. One nM leptin decreased the overall action potential frequency from 1.8 ± 0.3 Hz to 1.0 ± 0.2 Hz (p < 0.05), and 10 nM leptin decreased the overall action potential frequency from 0.9 ± 0.2 Hz to 0.6 ± 0.1 Hz (p < 0.05). However, 0.1 nM leptin did not significantly change the overall action potential frequency. In 1 mM extracellular Mg²⁺, 700 ms current steps of +20 pA, +30 pA, and +40 pA evoked 9 ± 1, 11 ± 1, and 13 ± 1 action potentials, respectively. Leptin (0.1, 1, and 10 nM) did not change the number of action potentials evoked by these steps. Leptin also did not alter action potential amplitude (107 ± 2 mV), duration at half-maximal amplitude (2.3 ± 0.2 ms), or threshold (-39 ± 2 mV). Leptin (0.1, 1, and 10 nM) did not change the resting membrane potential (-61 ± 2 mV) or input resistance (1030 ± 98 MΩ). Finally, leptin (0.1, 1 and 10 nM) did not alter the amplitude of the peak current evoked by 100 μM AMPA or 100 μM NMDA. Conclusions: These results indicate that leptin inhibits neuronal bursting in cultured hippocampal neurons induced by a Mg²⁺-free extracellular solution. They also suggest that this anticonvulsant effect does not result from a change in action potentials, resting membrane properties, or modulation of postsynaptic glutamate receptors. (Sources of funding – NIH, Epilepsy Foundation, Alafi Fund)