Fig. 6: Electrophysiological characterization of VMPOLepR neurons.
From: Thermally induced neuronal plasticity in the hypothalamus mediates heat tolerance
a, RMP in acclimated VMPOLepR neurons (n = 19/3) depolarized compared with the RMP of non-acclimated VMPOLepR (n = 17/3) cells. Unpaired, two-tailed Student’s t-test, ***P = 0.0001. b, Membrane input resistance (Rm) comparable between non-acclimated (n = 37/9) and acclimated (n = 41/10) VMPOLepR neurons. c, Left: membrane hyperpolarization in non-acclimated (n = 17/2) and acclimated (n = 19/2) VMPOLepR neurons caused by replacement Na+ for NMDG+ in aCSF. Right: the difference in membrane potential (Δ) between Na+-based aCSF and NMDG+-based aCSF is larger in acclimated VMPOLepR neurons. Unpaired, two-tailed Student’s t-test, *P = 0.0201. d, Left: AP phase plot of non-acclimated (gray, n = 9/4) and acclimated (red, n = 10/5) VMPOLepR neurons. Right: both AP 10–90% rise time (Wilcoxon’s test, **P = 0.0046) and 90% to 10% decay time (unpaired, two-tailed Student’s t-test, ***P = 0.0006) are significantly faster in VMPOLepR neurons after acclimation. e, Left: current–voltage relationship for VMPOLepR neuron peak transient NaV currents recorded in nucleated patches. Two-way ANOVA (effect of acclimation voltage, *P < 0.0001; Tukey’s multiple-comparison test, **P = 0.0016 (−25 mV), ***P = 0.0003 (−20 mV), ***P = 0.0002 (−15 mV), ***P < 0.0001 (−10 mV), ***P = 0.0005 (−5 mV) and **P = 0.0072 (0 mV); n = 6/2 (Non-accl.) and n = 6/2 (Accl.) cells). Right: example of transient NaV current recordings from VMPOLepR neurons. Inset: voltage step protocol used. f, Left: average INaP, revealed by slow depolarizing voltage ramp, enhanced after heat acclimation (n = 12/4 (Non-accl.) and n = 10/4 (Accl.) cells). Inset: ramp protocol used to record INaP. Right: quantification of INaP at −35 mV based on data shown on the left. Unpaired, two-tailed Student’s t-test, *P = 0.0055. g, Left: INaP in acclimated VMPOLepR neurons reduced by riluzole (10 µM) and completely blocked by TTX (1 µM). Right: quantification of INaP at −35 mV based on data shown on the left. One-way ANOVA, P < 0.0001; Tukey’s multiple-comparison test, ***P < 0.0001 (Accl.:Accl. + riluzole), ***P < 0.0001 (Accl.:Accl. + TTX), *P = 0.0170 (Accl. + riluzole:Accl. + TTX) (n = 9/2 (Accl.), n = 10/2 (riluzole) and n = 7/2 (TTX) cells). h, Left: firing frequency (fAP) of acclimated VMPOLepR neurons reduced by riluzole (10 µM) and ICA121431 (200 nM). One-way ANOVA, P < 0.0001; Tukey’s multiple-comparison test, ***P < 0.0001 (Accl.:Accl. + riluzole), ***P < 0.0001 (Accl.:Accl. + ICA121431); n = 40/10 (Accl.), n = 35/4 (riluzole) and n = 39/6 (ICA121431) cells. Right: example traces of the three conditions shown. i, Left: NaV1.3 antagonist ICA121341 blocking INaP in acclimated VMPOLepR neurons to a similar extent to riluzole. Right: quantification of INaP at −35 mV based on data shown on the left. One-way ANOVA, P = 0.0002; Tukey’s multiple-comparison test, **P = 0.0029 (Accl.:Accl. + ICA121431), ***P = 0.0001 (Accl.:Accl. + ICA121431 + riluzole); n = 8/3 (Accl.), n = 12/4 (ICA121431) and n = 10/2 (ICA121431 + riluzole). Part of the Accl. INaP data shown in g was repurposed for comparisons shown here. j, Distribution of temperature-insensitive, CSN, WSN and silent neurons within acclimated VMPOLepR neuron populations recorded with either riluzole (10 µM) or ICA121431 (200 nM) in perfusion fluid (n = 33/4 for riluzole and n = 24/4 for ICA121431). k, Firing frequencies of acclimated VMPOLepR control cells (n = 30/5), acclimated VMPOLepR cells recorded with riluzole (n = 33/4) and acclimated VMPOLepR cells recorded with ICA121431 (n = 24/4). Individual cells are plotted in color; black lines represent linear regression for each group Tcore (slope or temperature coefficient) = 1.9 for Accl. control, Tcore = 0.68 for riluzole and Tcore = 0.29 for ICA121431. Acclimated control cells were randomly sampled from the acclimated VMPOLepR cells plotted in Fig. 1e. Box plots in a–c and h represent the median and IQR; elsewhere data are shown as mean ± s.e.m (Extended Data Fig. 9 and Supplementary Figs. 1 and 2). Neuronal activity and currents were recoded under fast synaptic transmission blockade and at 36 °C.
