Phosphatidylinositol 4,5-bisphosphate (PIP₂ ) modulates afterhyperpolarizations in oxytocin neurons of the supraoptic...

Afterhyperpolarizations (AHPs) generated by repetitive action potentials in supraoptic magnocellular neurons regulate repetitive firing and spike frequency adaptation but relatively little is known about PIP₂ 's control of these AHPs. We examined how changes in PIP₂ levels affected AHPs, somatic [Ca²⁺ ]_i , and whole cell Ca²⁺ currents. Manipulations of PIP₂ levels affected both medium and slow AHP currents in oxytocin (OT) neurons of the supraoptic nucleus. Manipulations of PIP₂ levels did not modulate AHPs by influencing Ca²⁺ release from IP₃ -triggered Ca²⁺ stores, suggesting more direct modulation of channels by PIP₂ . PIP₂ depletion reduced spike-evoked Ca²⁺ entry and voltage-gated Ca²⁺ currents. PIP₂ appears to influence AHPs in OT neurons by reducing Ca²⁺ influx during spiking.

Oxytocin (OT)- and vasopressin (VP)-secreting magnocellular neurons of the supraoptic nucleus (SON) display calcium-dependent afterhyperpolarizations (AHPs) following a train of action potentials that are critical to shaping the firing patterns of these cells. Previous work demonstrated that the lipid phosphatidylinositol 4,5-bisphosphate (PIP₂ ) enabled the slow AHP component (sAHP) in cortical pyramidal neurons. We investigated whether this phenomenon occurred in OT and VP neurons of the SON. Using whole cell recordings in coronal hypothalamic slices from adult female rats, we demonstrated that inhibition of PIP₂ synthesis with wortmannin robustly blocked both the medium and slow AHP currents (I_mAHP and I_sAHP ) of OT, but not VP neurons with high affinity. We further tested this by introducing a water-soluble PIP₂ analogue (diC₈ -PIP₂ ) into neurons, which in OT neurons not only prevented wortmannin's inhibitory effect, but slowed rundown of the I_mAHP and I_sAHP . Inhibition of phospholipase C (PLC) with U73122 did not inhibit either I_mAHP or I_sAHP in OT neurons, consistent with wortmannin's effects not being due to reducing diacylglycerol (DAG) or IP₃ availability, i.e. PIP₂ modulation of AHPs is not likely to involve downstream Ca²⁺ release from inositol 1,4,5-trisphosphate (IP₃ )-triggered Ca²⁺ -store release, or channel modulation via DAG and protein kinase C (PKC). We found that wortmannin reduced [Ca²⁺ ]_i increase induced by spike trains in OT neurons, but had no effect on AHPs evoked by uncaging intracellular Ca²⁺ . Finally, wortmannin selectively reduced whole cell Ca²⁺ currents in OT neurons while leaving VP neurons unaffected. The results indicate that PIP₂ modulates both the I_mAHP and I_sAHP in OT neurons, most likely by controlling Ca²⁺ entry through voltage-gated Ca²⁺ channels opened during spike trains.