Supplementary Figure 6: Effects of THC on putative dopamine neurons of male offspring.
From: Prenatal THC exposure produces a hyperdopaminergic phenotype rescued by pregnenolone
(a) Representative traces show spontaneous dopamine firing activity recorded from both CTRL (ncells= 6, nrat= 6, nlitter= 3) and PCE (ncells= 7, nrat= 7, nlitter= 5) offspring before (basal) and after bath application of THC (3μM, 5 min). Calibration bar: 40 s, 50 mV. (b) Dose-response curves for the effects of THC on spontaneous dopamine cell activity (****P<0.0001, F(3,44)=8.96; two-way ANOVA) recorded from CTRL (ncells= 6, nrat= 6, nlitter= 3) and PCE (ncells= 7, nrat= 7, nlitter= 5). Data are represented as mean ± S.E.M. (c) THC effect is prevented by the CB1 antagonist AM281 (0.5 μM) in both CTRL (ncells= 6, nrat= 6, nlitter= 2) and PCE (ncells= 6, nrat= 6, nlitter= 2) progenies (interaction THC x PCE: P<0.0001, F(2,30)=20.33; two-way ANOVA) (d) Representative traces showing no effect of THC on AP waveform: AP waveform before (purple and light blue) and after THC (grey and blue) from CTRL (ncells= 6, nrat= 6, nlitter= 3) and PCE (ncells= 7, nrat= 7, nlitter= 5) dopamine cells. No alterations in amplitude (P=0.36, ns, t11=0.94; two-sided unpaired t-test) and rise time (P=0.24, ns, t11=1.236; two-sided unpaired t-test) were detected between CTRL and PCE offspring. Calibration bar: 10pA, 10ms. (e) Representative traces of action potentials (APs) elicited in response to the maximum current injected (200 pA) by dopamine neurons of CTRL (ncells= 7, nrat= 7, nlitter= 2) and PCE (ncells= 7, nrat= 7, nlitter= 5) offspring before (basal) and after bath application of THC (3 μM, 5 min). Calibration bar: 40mV, 300 ms. N= 7 experiments per group were repeated independently, with similar results obtained (f) THC net effect (Δvalue= number of AP THC- number of AP basal) on spike frequency in response to somatically injected current in CTRL and PCE offspring (**P=0.002, F(9,88)=3.22; Two-way RM ANOVA followed by Tukey’s multiple comparisons test; CTRL: ncells= 7, nrat= 7, nlitter= 2 and PCE: ncells= 7, nrat= 7, nlitter= 5. (g) Effect of THC on the latency to first AP appearance in response to the smallest current injected (50 pA) in PCE cells (*P=0.02, t(11)= 2.1; two-tailed unpaired t test). Top, Representative traces of evoked APs in response to the minimum current injected (50 pA) by dopamine neurons before (light blue trace) and after THC application (blue trace). Calibration bar: 1s, 20 mV. CTRL: ncells= 7, nrat= 7, nlitter= 2 and PCE: ncells= 7, nrat= 7, nlitter= 5. (h) Percentage of cells eliciting APs at 50 pA in CTRL and PCE animals in this set of experiments is consistent with those presented in Fig. 2g. Unless otherwise indicated, for all panels, graphs show box-and-whisker plots (including minima, maxima, and median values, and lower and upper quartiles) with single values expressing average data for each animal.
