Cocaine sensitization increases subthreshold activity in dopamine neurons from the ventral tegmental area

Abstract

The progressive escalation of psychomotor responses that results from repeated cocaine administration is termed sensitization. This phenomenon alters the intrinsic properties of dopamine (DA) neurons from the ventral tegmental area (VTA), leading to enhanced dopaminergic transmission in the mesocorticolimbic network. The mechanisms underlying this augmented excitation are nonetheless poorly understood. DA neurons display the hyperpolarization-activated, nonselective cation current, dubbed I_h We recently demonstrated that I_h and membrane capacitance are substantially reduced in VTA DA cells from cocaine-sensitized rats. The present study shows that 7 days of cocaine withdrawal did not normalize I_h and capacitance. In cells from cocaine-sensitized animals, the amplitude of excitatory synaptic potentials, at -70 mV, was ∼39% larger in contrast to controls. Raise and decay phases of the synaptic signal were faster under cocaine, a result associated with a reduced membrane time constant. Synaptic summation was paradoxically elevated by cocaine exposure, as it consisted of a significantly reduced summation indexed but a considerably increased depolarization. These effects are at least a consequence of the reduced capacitance. I_h attenuation is unlikely to explain such observations, since at -70 mV, no statistical differences exist in I_h or input resistance. The neuronal shrinkage associated with a diminished capacitance may help to understand two fundamental elements of drug addiction: incentive sensitization and negative emotional states. A reduced cell size may lead to substantial enhancement of cue-triggered bursting, which underlies drug craving and reward anticipation, whereas it could also result in DA depletion, as smaller neurons might express low levels of tyrosine hydroxylase.

New & noteworthy: This work uses a new approach that directly extracts important biophysical parameters from alpha function-evoked synaptic potentials. Two of these parameters are the cell membrane capacitance (C_m) and rate at any time point of the synaptic waveform. The use of such methodology shows that cocaine sensitization reduces C_m and increases the speed of synaptic signaling. Paradoxically, although synaptic potentials show a faster decay under cocaine their temporal summation is substantially elevated.

Keywords: EPSP; capacitance; cocaine; h-current; temporal summation.

Fig. 1.

Seven days of cocaine administration leads to the development of locomotor sensitization. All cocaine-treated rats used in this study expressed locomotor sensitization after a 7-day intraperitoneal injection (15 mg/kg) protocol. For each day, rats were first habituated to recording cages for 15 min, and data (means ± SE) were sampled every 5 min. Immediately after the conclusion of the habituation period, animals were injected either with saline solution (0.9%) or cocaine, and motor activity was recorded at 10 min intervals for 60 min. The motor activity measured at each time interval equals the sum of horizontal plus stereotype activities. Asterisks are for the comparison between days 1 and 7 of cocaine-treated rats: one-way ANOVA: F(3,24) = 79.14, ***P < 0.0001; post hoc comparison: Newman-Keuls multiple comparison test.

Fig. 2.

The cocaine-evoked reduction in I_h and C_m persists after 7 days of drug withdrawal. A₁: representative voltage-clamp traces showing the effects of cocaine sensitization onto I_h of VTA DA neurons following 7 days of drug withdrawal. As seen in the figure, I_h amplitude is defined as the difference between the steady-state current (I_ss) and the instantaneous current (I_ins). I_hold, holding current. A₂: summary graph of the records presented in A₁ showing that I_h amplitude reduction persists after 7 days of cocaine withdrawal. The direction of all depicted current amplitudes (Ampl.) is inward. B: I_h current inward rectification continues to be clearly depressed subsequent to 7 days of cocaine withdrawal. C: DA C_m continues to be significantly reduced after 7 days of cocaine withdrawal. C_m was determined in the I_h experiments summarized in B using the voltage step from −60 to −70 mV. The first 40 ms of the initial capacitive transient was fitted with a sum of 2 exponential functions, and the time constant of the fastest exponential term (τ_fast) was used to estimate C_m, according to the following expression: C_m = τ_fast(1/R_S + 1/R_N), where R_S is the series resistance, and R_N is the input resistance. D: summary graph illustrating that I_h current density is significantly reduced, 7 days after ending a cocaine-sensitization injection protocol. Current density was computed as the ratio of maximal I_h current (V_hold = −130 mV) to C_m. In summary plots A₂, C, and D, horizontal black segments with error bars represent means ± SE; unpaired t-test: *P < 0.05, **P < 0.01.

Fig. 3.

Cocaine sensitization augments the αEPSP amplitude. A₁: representative αEPSP recordings of VTA DA neurons from cocaine (gray trace)- and saline (black trace)-treated animals after a 7-day withdrawal period. αEPSPs were generated by the somatic injection of αEPSCs with α = 5 ms and I_max = 50 pA. Each record shown in the figure is the average of 10 consecutive sweeps. A₂: summary plot showing that cocaine sensitization leads to a significant increase in αEPSP amplitude. B: examples of least square fits of Eq. 2 to αEPSP recordings of cells from cocaine (gray trace with dark-gray-fitted curve)- and saline (black trace with gray-fitted curve)-treated animals. The use of the fitted coefficients allowed the estimation of R_N, C_m, and τ (τ_m; values for each shown in gray for cocaine and black for saline). C: summary graph showing that cocaine sensitization did not alter R_N at −70 mV. D: summary graph showing that cocaine sensitization resulted in a significant reduction in C_m. E: summary graph showing that cocaine sensitization resulted in a significant reduction in τ_m. In summary graphs A₂, C, D, and E, horizontal black segments with error bars represent means ± SE; unpaired t-test: *P < 0.05, **P < 0.01.

Fig. 4.

Cocaine sensitization augments the αEPSP rate (dV/dt). A₁ and A₂: representative examples illustrating the calculations of the temporal development of the dV/dt. Computations were performed with Eq. 3. Each αEPSP used in this analysis is the average of 10 consecutive sweeps. Dashed lines indicate that the peak αEPSP value happens exactly when dV/dt = 0. B: summary plot showing that cocaine sensitization leads to a significant augmentation of the MDR. C: summary plot showing that cocaine sensitization leads to a significant augmentation of the MRR. In summary plots B and C, horizontal black segments with error bars represent means ± SE; unpaired t-test: **P < 0.01.

Fig. 5.

Cocaine sensitization increases temporal summation at −70 mV. A: representative temporal summation recordings evoked by the somatic injection of a train of 5αEPSCs (α = 5 ms; I_peak = 50 pA) at 33 Hz (period: 30 ms) of VTA DA neurons from cocaine (gray trace)- and saline (black trace)-treated animals after a 7-day withdrawal period. Each record is the average of 10 consecutive sweeps. B: summary plot showing that cocaine sensitization leads to a significant augmentation of the first and fifth αEPSP at −70 mV. C: summary plot showing that cocaine sensitization leads to a significant reduction of the TSI at −70 mV. D: summary plot showing that cocaine sensitization leads to a significant augmentation of the mean depolarization endured during temporal summation at −70 mV. E: summary plot showing that cocaine sensitization did not alter the variability around the mean depolarization measure in D. In summary plots B, C, D, and E, horizontal black segments with error bars represent means ± SE; unpaired t-test: *P < 0.05, **P < 0.01. CVar, coefficient of variation.

Fig. 6.

Cocaine sensitization increases the membrane time constant of VTA DA cells at −100 mV. A: representative temporal summation recordings generated by the somatic injection of a train of 5αEPSC (α = 5 ms; I_peak = 50 pA) at 33 Hz of VTA DA neurons from cocaine (gray trace)- and saline (black trace)-treated animals after a 7-day withdrawal period. Each record is the average of 10 consecutive sweeps. The first αEPSP was least square fitted with Eq. 2. Cocaine: gray trace with black-fitted first αEPSP; saline: black trace with gray-fitted first αEPSP. B: summary graph showing that cocaine sensitization caused a significant augmentation of R_N at −100 mV. C: summary graph showing that cocaine sensitization resulted in a significant reduction in C_m at −100 mV. D: summary graph showing that cocaine sensitization resulted in a significant augmentation in τ at −100 mV. In summary graphs B, C, and D, horizontal black segments with error bars represent means ± SE; unpaired t-test: *P < 0.05, **P < 0.01.

Fig. 7.

Cocaine sensitization increases temporal summation at −100 mV. A: summary plot showing that cocaine sensitization leads to a significant augmentation of the TSI at −100 mV. B: summary plot showing that cocaine sensitization leads to a significant augmentation of the mean depolarization endured during temporal summation at −100 mV. C: summary plot showing that cocaine sensitization did not alter the variability around the mean depolarization measure in B. D₁ and D₂: overlay of representative temporal summation recordings of the same cell held at −70 and −100 mV. Each record is the average of 10 consecutive sweeps. Observe that cells from cocaine-sensitized rats, in contrast to saline controls, manifest and increase temporal summation at −100 mV. In summary plots A, B, and C, horizontal black segments with error bars represent means ± SE; unpaired t-test: *P < 0.05, **P < 0.01. CVar, coefficient of variation.