Comparison of electrophysiological properties of two types of pre-sympathetic neurons intermingled in the hypothalamic paraventricular nucleus

Abstract

The hypothalamic paraventricular nucleus (PVN) contains two types of neurons projecting to either the rostral ventrolateral medulla (PVN_RVLM) or the intermediolateral horn (IML) of the spinal cord (PVN_IML). These two neuron groups are intermingled in the same subdivisions of the PVN and differentially regulate sympathetic outflow. However, electrophysiological evidence supporting such functional differences is largely lacking. Herein, we compared the electrophysiological properties of these neurons by using patch-clamp and retrograde-tracing techniques. Most neurons (＞70%) in both groups spontaneously fired in the cell-attached mode. When compared to the PVN_IML neurons, the PVN_RVLM neurons had a lower firing rate and a more irregular firing pattern (p ＜ 0.05). The PVN_RVLM neurons showed smaller resting membrane potential, slower rise and decay times, and greater duration of spontaneous action potentials (p ＜ 0.05). The PVN_RVLM neurons received greater inhibitory synaptic inputs (frequency, p ＜ 0.05) with a shorter rise time (p ＜ 0.05). Taken together, the results indicate that the two pre-sympathetic neurons differ in their intrinsic and extrinsic electrophysiological properties, which may explain the lower firing activity of the PVN_RVLM neurons. The greater inhibitory synaptic inputs to the PVN_RVLM neurons also imply that these neurons have more integrative roles in regulation of sympathetic activity.

Keywords: action potential; inhibitory postsynaptic current; patch-clamp techniques; rostral ventrolateral medulla; spinal cord lateral horn.

Fig. 1. The firing activity in PVN_RVLM and PVN_IML neurons. (A) Representative traces showing spontaneous firing activities of a PVN_RVLM neuron and a PVN_IML neuron. (B and C) Cumulative bar graphs showing the mean frequency of spontaneous spikes (B) and the coefficient of variance (CV) of the interspike interval (ISI) in PVN_RVLM and PVN_IML neurons (C). (D) Plot of firing rate vs. the CV of the ISI. The curves were drawn by fitting the data with the equation y = a + b × c^x. The CV values in the PVN_IML neurons tended to be smaller than those in the PVN_RVLM neurons at the same firing rate. The values in the bars (B and C) represent the total number of neurons tested. Values represent mean ± SEM. Asterisks indicate p < 0.05 obtained by the unpaired Student's t-test. PVN, paraventricular nucleus; RVLM, rostral ventrolateral medulla; IML, intermediolateral horn of the spinal cord.

Fig. 2. The parameters of spontaneous action potentials in PVN_RVLM and PVN_IML neurons. (A) Representative action potentials from a PVN_RVLM neuron (solid line; RMP, −56.07 mV; threshold, −43.09 mV; rise time, 0.95 msec; decay time, 1.05 msec; half-APD, 2.15 msec) and a PVN_IML neuron (dotted line; RMP, −60.53 mV; threshold, −39.93 mV; rise time, 0.6 msec; decay time, 0.75 msec; half-APD, 1.65 msec) were overlapped at threshold. Cumulative bar graphs showing the 10% to 90% rise time (B), 37% to 90% decay time (C) and half-APD (D) in PVN_RVLM and PVN_IML neurons. The values in the bars (B–D) represent the total number of neurons tested. Values represent mean ± SEM. Asterisks indicate p < 0.05 by the unpaired Student's t-test. PVN, paraventricular nucleus; RVLM, rostral ventrolateral medulla; IML, intermediolateral horn of the spinal cord; half-APD, duration of action potential at half amplitude.

Fig. 3. Miniature inhibitory postsynaptic current (mIPSC) in PVN_RVLM and PVN_IML neurons. Representative current traces of mIPSCs in a PVN_RVLM neuron (A) and a PVN_IML neuron (B) (V_h = −70 mV). Cumulative bar graphs showing the mean frequency (C) and mean amplitude (D) of the mIPSCs. The values in the bars (C and D) represent the total numbers of neurons tested. Values are mean ± SEM. Asterisk indicates p < 0.05 by the unpaired Student's t-test. PVN, paraventricular nucleus; RVLM, rostral ventrolateral medulla; IML, intermediolateral horn of the spinal cord.

Fig. 4. The miniature inhibitory postsynaptic current (mIPSC) kinetics in the PVN_RVLM and the PVN_IML neurons. (A) Representative traces of mIPSCs in a PVN_RVLM neuron (black) and a PVN_IML neuron (gray) (left). Illustration of the 10% to 90% rise time and the decay time at an expanded time scale (middle and right). (B) Summary bar graphs showing the mean 10% to 90% rise time in the PVN_RVLM and PVN_IML neurons. (C) Summary bar graphs displaying the weighted decay time constant under a well-fitted second exponential. The values in the bars (B and C) represent the total numbers of neurons tested. Values are mean ± SEM. Double asterisk indicates p < 0.05 by the unpaired Student's t-test. PVN, paraventricular nucleus; RVLM, rostral ventrolateral medulla; IML, intermediolateral horn of the spinal cord.