TC-2559 excites dopaminergic neurones in the ventral tegmental area by stimulating alpha4beta2-like nicotinic acetylcholine receptors in anaesthetised rats

Abstract

1. The in vivo effects of a selective partial agonist for neuronal nicotinic acetylcholine receptor (nAChRs) alpha4beta2 subtype, TC-2559, characterised recently in in vitro preparations, have been profiled. The brain bioavailability of TC-2559 and its effects on the spontaneous firing and bursting properties of the dopaminergic (DAergic) neurones recorded extracellularly in the ventral tegmental area (VTA) were studied following systemic administration in anaesthetised rats. 2. Cumulative doses of TC-2559 (0.021-1.36 mg kg(-1), i.v.) increased both the firing and bursting activities of VTA DA neurones. The effect of bolus doses of TC-2559 of 0.66 or 1.32 mg kg(-1), i.v., was approximately equivalent to that of 0.0665 mg kg(-1), i.v. nicotine. 3. The excitation evoked by both nicotine and TC-2559 was fully reversed by DHbetaE (0.39-0.77 mg kg(-1), i.v.), an alpha4beta2-subtype-preferring nicotinic antagonist, and application of nicotine after DHbetaE failed to evoke any excitation. MLA (0.23 mg kg(-1), i.v.), an alpha7 selective antagonist, failed to alter TC-2559-evoked excitation and bursting activities, and a novel alpha7 agonist (PSAB-OFP; 0.23 mg kg(-1), i.v.) was also without effect. 4. The present results indicated that TC-2559 fully mimics nicotine by increasing both the excitability and bursting behaviour of VTA DA neurones, effects that are predominantly due to activation of alpha4beta2-like nAChRs. 5. TC-2559 has been demonstrated to be a useful in vivo pharmacological tool for studying the alpha4beta2 subtype of nicotinic receptor.

Figure 1

Traces showing total brain concentration of TC-2559 (1.32 mg kg⁻¹, i.v.), DHβE (0.77 mg kg⁻¹, i.v.) and PSOB-OFP (0.23 mg kg⁻¹, i.v.) from 1.5 to 30 min after i.v. bolus injection. (Each data point is an average from two animals.)

Figure 2

Effect of nicotine and subsequent doses of DHβE on the firing and bursting rates of a VTA DA neurone in choral hydrate anaesthetised rat. (a) Rate histogram showing that bolus dose of nicotine (0.0665 mg kg⁻¹) increased neuronal firing. Arrows indicate the time when saline, nicotine or DHβE was injected intravenously. ‘i–iv' above the rate histogram indicate the periods when the inter-spike interval histograms (ISIHs) in (b) were recorded. (b) ISIH showing nicotine also increased bursting activity. (i) control; (ii, iii) after nicotine injection; (iv) after DHβE injection. Both the rate and bursting change evoked by nicotine were reversed by DHβE (0.77 mg kg⁻¹, i.v.). (In (b), f=firing frequency, B=% of spikes in burst.)

Figure 3

Effect of cumulative doses of TC-2559 and subsequent doses of DHβE and nicotine on the firing and bursting rates of a VTA DA neurone in choral hydrate anaesthetised rat. (a) Rate histogram showing that cumulative doses of TC-2559 (0.085, 0.17, 0.34, 0.68 and 1.36 mg kg⁻¹, intravenously at arrows) dose dependently increased neuronal firing. ‘i–vi' above the rate histogram indicate the periods (3 min) when the ISIHs in (b) were recorded. (b) ISIHs showing cumulative doses of TC-2559 also increased the bursting activity. (i) Control; (ii) after saline injection, (iii, iv) after TC-2559 injection; (v) after DHβE injection and (vi) after nicotine injection. Both the rate and bursting changes evoked by TC-2559 were reversed by a single dose of DHβE (0.77 mg kg⁻¹, i.v.). Subsequent application of nicotine failed to induce changes in either rate or bursting. (c) Group data showing TC-2559 significantly (^*P<0.05) increased the VTA DA neurone activity, which was subsequently reversed by DHβE (^#P<0.05). Neuronal activity inhibited by apomorphine (17–52 μg kg⁻¹ (a, c)) confirmed that the recorded cells were DAergic neurones. (In (b), f=firing frequency, B=% of spikes in burst.)

Figure 4

Effect of TC-2559 and subsequent doses of DHβE, nicotine and apomorphine on the firing and bursting rates of a VTA DA neurone in choral hydrate anaesthetised rat. (a) Rate histogram showing that bolus dose of TC-2559 (0.66 mg kg⁻¹) increased neuronal firing. Arrows indicate the time when saline or drugs were injected intravenously. ‘i–vi' above the rate histogram indicate the periods (3 min) when the ISIHs in (b) were recorded. (b) ISIH showing TC-2559 also increased the bursting activity. (i) Control; (ii) after saline; (iii, iv) after TC-2559; (v) after DHβE; and (vi) after nicotine injection. Both the rate and bursting changes evoked by TC-2559 were reversed by a single dose of DHβE (0.77 mg kg⁻¹, i.v.). Subsequent application of nicotine (0.0665 mg kg⁻¹) failed to induce changes in either rate or bursting. Neuronal activity inhibited by apomorphine (17 μg kg⁻¹, i.v.; (a) confirmed that the recorded cell was DAergic neurone). (In (b), f=firing frequency, B=% of spikes in burst.)

Figure 5

Effect of MLA on the excitatory action of TC-2559 on the firing and bursting rates of a VTA DA neurone in choral hydrate anaesthetised rat. (a) Rate histogram showing with MLA (0.23 mg kg⁻¹) pre-treatment, bolus dose of TC-2559 (1.32 mg kg⁻¹) increased neuronal firing. Arrows indicate the time when saline, MLA, TC-2559 and DHβE was injected intravenously (note the long-lasting effect, >10 min, of the TC-2559). ‘i–v' above the rate histogram indicate the periods (3 min) when the ISIHs in (b) were recorded. (b) ISIHs showing TC-2559 also increased the bursting activity, despite the pre-treatment of the MLA. (i) saline injection; (ii) after MLA injection; (iii, iv) after TC-2559 injection; (v) after DHβE injection. Both the rate and bursting changes evoked by TC-2559 were reversed by a single dose of DHβE (0.77 mg kg⁻¹). Neuronal activity inhibited by apomorphine (13 × 3 μg kg⁻¹ (a)) confirmed that the recorded cell was DAergic neurone. (In (b), f=firing frequency, B=% of spikes in burst.)

Figure 6

Group data of the effects of single doses of drugs on the firing rate (a) and the bursting rate (b) of the VTA DA neurones. Bar histograms represent the group data (mean±s.e.m.) of nicotine (0.0665 mg kg⁻¹, n=15 in (a) and n=8 in (b)), TC-2559 (0.66 mg kg⁻¹, n=14 in (a) and n=12 in (b)) and TC-2559 (1.32 mg kg⁻¹, n=12 in (a) and n=9 in (b)). Statistics: ^*P<0.05 compared with saline control, and ^#P<0.05 of the fifth minute data after drug injection compared with the data taken after DHβE (0.77 mg kg⁻¹) injection.

Figure 7

Effect of PSAB-OFP and subsequent doses of nicotine on the firing and bursting rates of a VTA DA neurone. (a) Rate histogram showing that bolus dose of PSAB-OFP (0.23 mg kg⁻¹) had no effect on firing rate, but nicotine (0.0665 mg kg⁻¹) increased neuronal firing. Arrows indicate the time when saline, PSAB-OFP or nicotine was injected intravenously. ‘i–iv' above the rate histogram indicate the periods (3 min) when the inter spike interval histograms (ISIHs) in (b) were recorded. (b) ISIHs showing that PSAB-OFP had no effect but nicotine increased the bursting activity. (i) Control; (ii, iii) after PSAB-OFP injection; (iv) after nicotine injection. (In (b), f=firing frequency, B=% of spikes in burst.)

Figure 8

Effects of iontophoretic application of TC-2559 on the VTA DA neurone activity. Rate histogram showing that: (a) TC-2559 (10-40 nA) dose-dependently increased the firing rate and (b) TC-2559-evoked excitation was attenuated by co-application of DHβE (20 nA). The bars above the rate histogram indicate the time when drugs were ejected by iontophoresis.