Oxytocin modulates GABAA receptor-mediated inhibition onto CA1 pyramidal neurons in mouse

Oxytocin (OT) is a neuropeptide that exerts different peripheral and central actions. I aimed at characterizing the neuromodulatory effects of OT in the hippocampus. Electrophysiological experiments were performed on mouse brain slices using the whole-cell patch-clamp technique on pyramidal neurons (PYR) and GABAergic interneurons (INs) located in CA1 stratum pyramidale. The effect of TGOT (Thr4,Gly7-oxytocin), a selective OT receptor (OTR) agonist, was first evaluated on spontaneous inhibitory postsynaptic currents (sIPSC) recorded from PYRs in Otr+/+ mice. TGOT caused a significant decrease in the sIPSC interval and an increase in the sIPSC amplitude; it also caused an increase in the sIPSC time constant of decay: this suggests the involvement of GABAA receptors (GABAAR) located in a perisynaptic position that deactivate slower than synaptic receptors, generating slower sIPSCs. The TGOT-mediated effects were dependent on the activation of OTRs, being abolished by the OTR antagonist SSR126768A; furthermore, TGOT didn’t modulate sIPSCs in Otr-/- mice. Then, we recorded the miniature inhibitory postsynaptic currents (mIPSC), isolated by applying tetrodotoxin, a voltage-gated Na+ channel blocker, to prevent action potential firing in the presynaptic terminal. TGOT was not able to modulate the mIPSC interval, amplitude and kinetics of decay, indicating that the effects elicited by the agonist are dependent on the firing activity of the presynaptic neuron. After having clarified the action of TGOT on ‘phasic’ inhibitory transmission, elicited by synaptic and perisynaptic GABAARs, we enquire if the peptide could influence ‘tonic’ currents, mediated by extrasynaptic GABAARs. First, we demonstrated the presence of tonic currents by measuring the ‘baseline holding current’ required to clamp PYRs at a given potential, in control conditions and during the application of the GABAAR antagonist bicuculline: we observed an inward shift in the ‘baseline holding current’ in the presence of bicuculline, consistent with the abolition of tonic currents. Then, we found that TGOT was able to increase tonic currents, causing an outward shift in the ‘baseline holding current’. Subsequently, we tried to understand the source of that TGOT-mediated increased inhibition, finding that TGOT depolarized mainly the stuttering fast-spiking INs. The same depolarization was observed in the presence of synaptic blockers, suggesting that the effect is due to a direct binding to OTRs. Indeed, the perfusion of the OTR antagonist completely abolished the depolarization. We tried to investigate the ionic mechanism underlying the TGOT-induced depolarization. We tested the putative involvement of a Ca2+ current by using nifedipine, a selective L-type channel blocker. Actually, in the majority of INs examined, nifedipine was able to abolish the depolarization elicited by TGOT. Finally, we investigated the effect of TGOT on the membrane potential of PYRs. Most of them, examined at their spike threshold, became hyperpolarized by TGOT and their firing rate was significantly decreased. The hyperpolarizing response was completely abolished by the blockade of GABAARs, indicating that the effect requires the activation of extrasynaptic GABAARs that mediate a prolonged (or tonic) hyperpolarizing current. The TGOT-mediated hyperpolarization caused a reduction in cell excitability, altering the capability of PYRs to generate action potentials in response to depolarizing current steps. This was evident in the firing rate-to-injected current (F-I) relationship that was shifted to the right during perfusion of TGOT. The gain (i.e., the slope) of the curve was not influenced by TGOT. This behavior indicates an increase in tonically active inhibitory currents that lead to a persistent reduction in the input resistance and therefore in cell excitability.