Nicotine modulates GABAergic transmission to dopaminergic neurons in substantia nigra pars compacta

Abstract

Aim: Dopaminergic neurons in the substantia nigra pars compacta (SNc) play important roles in motor control and drug addiction. As the major afferent, GABAergic innervation controls the activity of SNc dopaminergic neurons. Although it is clear that nicotine modulates SNc dopaminergic neurons by activating subtypes of somatodendritic nicotinic acetylcholine receptors (nAChRs), the detailed mechanisms of this activation remain to be addressed.

Methods: In the current study, we recorded GABA(A) receptor-mediated spontaneous inhibitory postsynaptic currents (sIPSCs) from dissociated SNc dopaminergic neurons that were obtained using an enzyme-free procedure. These neurons preserved some functional terminals after isolation, including those that release GABA.

Results: We found that both extra- and intra-cellular calcium modulates sIPSCs in these neurons. Furthermore, both nicotine and endogenous acetylcholine enhance the frequency of sIPSCs. Moreover, endogenous acetylcholine tonically facilitates sIPSC frequency, primarily by activating the alpha4beta2* nAChRs on the GABAergic terminals.

Conclusion: Nicotine facilitates GABA release onto SNc dopaminergic neurons mainly via the activation of presynaptic alpha4beta2* nAChRs.

Figure 1

SNc DA neurons have distinct electrophysiological and pharmacological properties. A, spontaneous firing of a dissociated putative SNc DA neuron, which was reversibly depressed by 0.2 μmol/L quinpirole(QP) (Upper panels: typical traces; Lower panel: time course). B, under voltage clamp mode, an Ih current was induced using a series of incremental 10 mV hyperpolarizing steps from a holding potential of -60 mV.

Figure 2

Spontaneous GABA_A receptor-mediated IPSCs (sIPSCs) in DA neurons mechanically dissociated from the SNc. In the presence of 50 μmol/L APV and 20 μmol/L DNQX and at a holding potential (V_H) of -55 mV, sIPSCs were completely but reversibly abolished by bicuculline (Bic, 10 μmol/L) and were reversibly and significantly inhibited by TTX (1 μmol/L) and by CdCl₂ (50 μmol/L). Summary data (means±SEM) shows the effects of TTX, CdCl₂ and calcium free (Ca²⁺ free) on the frequency (upper panel) and amplitude of sIPSCs (lower panel). ^cP<0.01 compared to control.

Figure 3

Effects of nicotinic agonists on sIPSCs in SNc DA neurons. Typical traces of sIPSCs recorded under the control condition (A) and in the presence of 10 mmol/L choline (B), 1 mol/L nicotine (C), and 100 μmol/L RJR-2403 (D). All traces were from the same SNc DA neuron (V_H=-55 mV). EG: Summary of the effects of choline (E), nicotine (F), and RJR-2403 (G) on both frequency and amplitude of sIPSCs. ^bP<0.05, ^cP<0.01 compared to control.

Figure 4

Eserine enhances sIPSCs in SNc DA neurons. A-B, Eserine increases the frequency (Freq), but not amplitude (Ampl) of sIPSCs (A, typical traces; B, summary). C1 and C2, Eserine causes a leftward shift of the cumulative probability of interevent intervals of two consecutive sIPSCs (C1, K–S test, P<0.001), but not that of sIPSC amplitude (C2, K–S test, P= 0.13).

Figure 5

Effects of nicotinic antagonists on sIPSCs in SNc DA neurons. Representative traces of sIPSCs recorded under control conditions (A), in the presence of 300 nmol/L -BTX (B), 100 nmol/L DHβE (C), 10 μmol/L MEC (D), and after 10 min washout (E) with the standard external solution. All traces were from the same DA neuron (V_H=-55 mV). F: Cumulative probability plots show that DHE and MEC significantly increased incidence of long sIPSC interevent intervals (K–S test, P<0.001), but that α-BTX did not significantly alter the distribution of sIPSC interevent intervals (K–S test, P>0.05). G: Summary of the effects of α-BTX, DHβE, MEC on the frequency of sIPSCs. ^cP<0.01 compared to control.