Abstracts

Rationale: TNF-alpha is a critical mediator of neuro-inflammation and cellular loss or injury following brain insults, but the neonatal brain responds differently than the adult to pro-inflammatory cytokines. We previously reported increased TNF-alpha in neonatal rat hippocampus several days after hypoxia-induced seizures in the absence of cell loss or salient anatomical injury. In the current study, we aimed to determine if TNF-alpha could be detrimental by altering neuronal physiological activity. Methods: Long-Evans rat pups were subjected to 5-7% O2 for 15 min at age P10 to induce seizures. Whole-cell patch-clamp recordings were obtained from CA1 pyramidal neurons in acute hippocampal slices at P13 (3 days post-hypoxia). Current-clamp recordings of spontaneous activity were obtained for 10 minutes, TNF-alpha (0.5 ug/ml) was added to the bath, and recordings were continued for 30-40 minutes. For some cells, voltage-clamp recordings were obtained to examine TNF-alpha effects on intrinsic voltage-dependent currents. Results: TNF-alpha had little effect on resting membrane potential (RMP) or intrinsic membrane properties of the majority of CA1 pyramidal neurons in slices from control animals, but 2/10 (20%) of cells showed a depolarization of the RMP and increase in spontaneous firing that developed within 10-15 minutes of TNF-alpha application. In contrast, 8/11 (73%) of neurons in hypoxia-treated group exhibited RMP depolarization and increased spontaneous firing in response to TNF-alpha (p = 0.030, Fisher s Exact test). No significant differences were observed in RMP between groups prior to TNF-alpha application. Voltage-clamp recordings after TNF-alpha indicated a decrease in depolarization-activated outward currents in neurons from the hypoxia-treated group compared to controls, suggesting that decreased voltage-activated K currents could have mediated or contributed to the excitatory response. Conclusions: Hippocampal CA1 pyramidal neurons exhibited an augmented excitatory response to TNF-alpha after neonatal hypoxia-induced seizures sub-acutely and at a time when TNF-alpha levels are elevated. This increased excitation could promote repeated seizures after the initial insult, and potentially contribute to AED refractoriness and exacerbate long-term adverse outcomes.