Fig. 2.

Examples of actions of picospritzed HA and the rapid E2 effects under “K⁺-only” condition. The figure shows two sets of 3 segments of continuous whole-cell recordings of membrane potential from two VMN neurons, A and B. The recordings were made in K⁺-only situation (A, in TTX plus Ca⁺⁺-free; and B, Na⁺- and Ca⁺⁺-free ACSFs; and K⁺-only internal solution). HA (10 mM in ejecting pipets containing respective K⁺-only ACSF) was picospritzed (for 2–3 sec) repetitively before, during and after acute bath application of E2 (50 nM). To facilitate comparisons, the horizontal dotted lines were drown aligning the Vm (A, −69.6 mV; B, −58.2 mV) at the point of each HA ejection. In A, HA depolarization was potentiated by E2 to 141%, from 2.44 to 3.45 mV. Neuron B responded biphasically to HA. The biphasic response was probably, as we have shown in a previous study [35], a manifestation of the colocalization of HA receptor subtypes H2/H3 and H1, which mediate hyper- and de-polarization, respectively. E2 rapidly potentiated the depolarization from 1.17 mV to 1.55 mV (or 132%) and abolished the hyperpolarization. Note, in B, the HA depolarization at HA+E2 sustained longer than those before and after E2 application. Also note that the membrane potential of each neuron remained stable despite the application of E2.