Authors :
Presenting Author: Andrew Huang, PhD Candidate – University of California Riverside
Deepak Subramanian, PhD – University of California - Riverside
Carol Eisenberg, PhD – Rutgers University
Michael W. Shiflett, PhD – Rutgers University
Tracy S. Tran, PhD – Rutgers University
Viji Santhakumar, PhD – University of California, Riverside
Rationale:
Polymorphisms in Neuropilin-2 (Nrp2), a semaphorin receptor critical for circuit formation and synapse maintenance, are clinically linked to autism spectrum disorders. Global Nrp2 deletion or selective embryonic deletion in interneuron precursors leads to excitation-inhibition imbalance, behavioral deficits, and seizure susceptibility, consistent with the features of autism-epilepsy phenotype. However, the effects of postnatal deletion of Nrp2 to circuit function and behaviors remain unclear. This study investigated the cellular, synaptic, network and behavioral consequences of postnatal Nrp2 deletion in the neocortex.
Methods:
Nrp2f/f::Etv1-CreERT2 mice and Cre-negative controls were administered tamoxifen at P7-8 to selectively delete Nrp2 in layer 5 pyramidal neurons, generating excitatory conditional knockout (eCKO) mice. Whole-cell patch clamp recordings were performed on Layer 5 pyramidal neurons to assess intrinsic physiology and miniature excitatory and inhibitory postsynaptic currents (mEPSC/mIPSC). Seizure susceptibility was evaluated as the latency to convulsive seizures following kainic acid injection (20mg/kg; i.p.). Mice were monitored up to 75 minutes post induction. Novel Object Recognition and Social Novelty tasks were assessed for proportion of time spent investigating the novel object or mouse, respectively. Results:
Layer 5 pyramidal neurons from eCKO mice showed reduced firing frequency in response to current injections (p=0.0175, +40-600pA, 2-way ANOVA). Analysis of mEPSCs revealed an increase in interevent interval (control: 0.21±0.019, eCKO: 0.38±0.023, n = 7 and 8 cells, p< 0.0001, KS test) and amplitude (control: 8.91±0.15, eCKO: 9.70±0.13, n = 7 and 8 cells, p< 0.0001, KS test) in eCKO mice. Similarly, mIPSC interevent interval and amplitude were also elevated in eCKO mice. While 2 of 5 eCKO mice exhibited spontaneous electrographic seizures, latency to kainic-acid induced seizures did not reach statistical significance (control: 21.47±3.97 min, eCKO: 13.28±3.05 min, n = 5 mice per group, p=0.14, t-test). However, eCKO mice exhibited higher mortality (3/5) from evoked seizures than controls (0/5). In the novel object recognition task, eCKO mice spent less time investigating the novel object (p=0.036, ANOVA) and allocated significantly less time with the novel mouse (p=0.02, ANOVA) in a social novelty preference task.