Abstracts

Rationale: The anticonvulsant effect of pharmacological inhibition of the substantia nigra pars reticulara (SNpr), originally identified in the early 1980s, has long been hypothesized to function by disinhibition of the deep and intermediate layers of the superior colliculus (SC). Consistent with this pharmacological or optogenetic activation of the SC is robustly anticonvulsant across models. However, the SNpr projects to other structures including the pedunculopontine nucleus (PPN). Pharmacological activation of the PPN has produced mixed effects across studies, with some reporting anticonvulsant effects. Selective targeting of nigral projections (i.e., SN-SC vs SN-PPN) was not attainable using pharmacological approaches. However, optogenetic methods now enable the direct targeting of these efferent pathways, and thus a direct test of these specific projections are now possible. Methods: Rats were injected with 1.5 uL of AAV coding for either inhibitory opsin, SwiChR or ArchT, in the SNpr and fiber optics were placed in the SNpr (to silence cell bodies). We compared these animals to those in which AAV-ArchT was injected into the SNpr and fiber optics were placed within the SC or PPN, to silence nigrotectal or nigrotegmental terminals. We compared the efficacy of optogenetic silencing across several models of experimentally-evoked seizures: systemic pentylenetetrazole (generalized seizures), focal microinjection of bicuculline into the “area tempestas” (focal limbic seizures), and systemic gamma butyrolactone (GBL; absence-like seizures). Results: Inhibition of cell bodies within the SNpr supressed seizures in all model examined. Optogenetic inhibition of the nigral cell bodies decreased the duration of time seizing after administration of GBL (n=9, p<0.005) with GBL. Optogenetic silencing of nigral cell bodies also decreased seizure score with both systemic PTZ (n=8, p<0.05) and focal microinjection of bicuculline (n=9, p<0.005). A similar profile of anticonvulsant action was seen after silencing nigrotectal (SN-SC) projections. Silencing nigral terminals in the SC decreased the  duration of time seizing (n=8, p<0.05) following GBL. Optogenetic silencing of nigral cell terminals in the SC also decreased seizure score with both systemic PTZ (n=10 p<0.005) and focal microinjection of bicuculline (n=9, p=0.005).  However, silencing SN-PPN projections produced a mixed effect. Inhibition of SN-PPN suppressed absence seizures (n=7, p=0.05) with GBL. Optogenetic silencing of nigral terminals in the PPN had no effect on seizure score with focal microinjection of bicuculline (n=5, p=0.5) and it worsened seizures evoked by PTZ (n=7, p=0.03).  Conclusions: Selective suppression of nigral terminals in the SC, but not PPN is sufficient to account for the full spectrum of anticonvulsant effects achieved by inhibition of the SNpr. These data indicate a functional divergence in this seizure control pathway, as silencing either SN-SC or SN-PPN projections disrupted absence seizures, but only silencing of nigrotectal projections disrupted limbic motor seizures. Funding: 5R01NS097762