Abstracts

Rationale: In the neonatal transcranial freeze lesion model of 4-layered microgyria, we have previously shown that Somatostatin (SS) and Parvalbumin interneurons are differentially affected within and around the microgyrus (paramicrogyral region, PMR).  Here we use optogenetics to study the selective output from specific interneuron subtypes.  We hypothesize that in malformed cortex hyperactive SS contribute to epileptiform activity via synchronization of columnar activity or network dis-inhibition through their synaptic contacts onto other inhibitory interneurons.  Methods: Whole cell patch clamp recordings were made in ex vivo slices from layer V pyramidal neurons within the PMR. IPSCs were recorded in isolation with glutamate antagonists APV and DNQX in the bath, a high K+ intracellular solution (ECl-  = -15 mV), and a voltage clamp holding potential of -70 mV.  Blue light was applied through a 60X objective centered over the recorded neuron in order to activate the Channelrhodopsin and depolarize SS interneurons. Results: An intensity series created by varying light duration showed increased peak responses in PMR compared to control cortex.  Recordings were also made from SS interneurons demonstrating that the number of action potentials generated in response to light activation was not different in PMR vs control cortex.  Instead repetitive light activation suggested an increased release probability. When the light application was moved to layer III above the recorded neuron, there was no longer a significant difference in the IPSC peak, suggesting a selective layer V locus. An intensity series of electrical stimuli was applied ~0.1 mm lateral to the recorded neuron before and during simultaneous light application.  In control cortex, adding light activation of SS interneurons did not significantly change the amplitude of the electrically evoked IPSC.  In contrast in the PMR, the addition of the increased SS activation decreased the amplitude of the electrically evoked IPSC.  This likely reflects an increased synaptic effectiveness (either in number or synaptic strength) of SS interneurons onto other inhibitory interneurons within the PMR.  Such an increase is likely to also reduce the effectiveness of Parvalbumin and other inhibitory interneuron types to effectively control intracortical excitation.  This idea is supported by our recordings of field potentials.  Under conditions of reduced inhibition (0.2 mM Gabazine in bathing medium) or increased excitation (low magnesium bathing aCSF), light activation of SS interneurons can evoke epileptiform activity in the PMR.  The amount of epileptiform activity evoked under these conditions is greater in the PMR than in control cortex. Conclusions: Together with our previous results, these findings suggest that the role of SS interneurons is increased in the PMR, with an increased excitatory input, an increased rate of intrinsic firing, an increased output likely due to increased release probability, and in increased effectiveness in depressing other inhibition, resulting in epileptiform activity. Funding: Supported by NIH NS054210.