Dopamine transporter down-regulation following repeated cocaine: implications for 3,4-methylenedioxymethamphetamine-induced acute effects and long-term neurotoxicity in mice

Abstract

Background and purpose: 3,4-Methylenedioxymethamphetamine (MDMA) and cocaine are two widely abused psychostimulant drugs targeting the dopamine transporter (DAT). DAT availability regulates dopamine neurotransmission and uptake of MDMA-derived neurotoxic metabolites. We aimed to determine the effect of cocaine pre-exposure on the acute and long-term effects of MDMA in mice.

Experimental approach: Mice received a course of cocaine (20 mg*kg(-1), x2 for 3 days) followed by MDMA (20 mg*kg(-1), x2, 3 h apart). Locomotor activity, extracellular dopamine levels and dopaminergic neurotoxicity were determined. Furthermore, following the course of cocaine, DAT density in striatal plasma membrane and endosome fractions was measured.

Key results: Four days after the course of cocaine, challenge with MDMA attenuated the MDMA-induced striatal dopaminergic neurotoxicity. Co-administration of the protein kinase C (PKC) inhibitor NPC 15437 prevented cocaine protection. At the same time, after the course of cocaine, DAT density was reduced in the plasma membrane and increased in the endosome fraction, and this effect was prevented by NPC 15437. The course of cocaine potentiated the MDMA-induced increase in extracellular dopamine and locomotor activity, following challenge 4 days later, compared with those pretreated with saline.

Conclusions and implications: Repeated cocaine treatment followed by withdrawal protected against MDMA-induced dopaminergic neurotoxicity by internalizing DAT via a mechanism which may involve PKC. Furthermore, repeated cocaine followed by withdrawal induced behavioural and neurochemical sensitization to MDMA, measures which could be indicative of increased rewarding effects of MDMA.

Figure 1

Effect of pretreatment with repeated cocaine (20 mg·kg⁻¹, i.p. twice daily separated by 8 h for 3 days) finishing 1 or 4 days before 3,4-methylenedioxymethamphetamine (MDMA) (20 mg·kg⁻¹, i.p., two injections separated 3 h) on (A) rectal temperature immediately following MDMA administration and (B) [³H]WIN 35 428 binding. Data shown as mean ± SEM, n= 5 − 11. There were no differences between the data obtained from saline controls pretreated 1 or 4 days earlier with cocaine and therefore data were grouped. (A) The MDMA-induced increase in rectal temperature (F_1,11= 54.79; P < 0.001) was not modified by pretreatment with cocaine 1 day (F_1,13= 0.97; P= 0.32, n.s.) or 4 days (F_1,11= 0.93; P= 0.33, n.s.) before. Cocaine pretreatment did not modify the temperature of saline-treated animals (F_1,15= 3.64; P= 0.06, n.s.). Two-way analysis of variance (anova). (B) Different from saline: *P < 0.001; different from MDMA: ^ΔP < 0.001; different from cocaine + MDMA 1 day later: ^δP < 0.001; different from cocaine + saline: ^fP < 0.05. One-way anova followed by Newman–Keuls test.

Figure 2

Effect of co-administration of NPC 15437 (1 mg·kg⁻¹, i.p.) and cocaine (20 mg·kg⁻¹, i.p. twice daily separated by 8 h for 3 days) finishing 4 days before 3,4-methylenedioxymethamphetamine (MDMA) (20 mg·kg⁻¹, i.p., two injections separated 3 h) on striatal (A) [³H]WIN 35 428 binding and (B) dopamine (DA) concentration, 7 days later. Data shown as mean ± SEM, n= 6 − 11. Different from saline *P < 0.001; different from MDMA: ^ΔP < 0.001; different from cocaine + MDMA: ^δP < 0.05, ^δδδP < 0.001; different from NPC (during cocaine treatment + withdrawal) + cocaine + MDMA: ^fP < 0.01, ^ffP < 0.001. One-way anova followed by Newman–Keuls test.

Figure 3

Effect of repeated cocaine (20 mg·kg⁻¹, i.p. twice daily separated by 8 h for 3 days) on striatal plasma membrane (A) and endosome (B) [³H]WIN 35 428 binding 1 or 4 days later. A group of animals were co-administered NPC 15437 (1 mg·kg⁻¹, i.p.) with cocaine and twice daily during the 3-day withdrawal. Data shown as mean ± SEM, n= 7 − 10. Different from saline: *P < 0.01; different from cocaine 1 day before: ^fP < 0.05, ^ffP < 0.01; different from cocaine only 4 days before: ^δP < 0.05. One-way anova followed by Newman–Keuls test.

Figure 4

Effect of pretreatment with repeated cocaine (20 mg·kg⁻¹, i.p. twice daily separated by 8 h for 3 days) finishing 1 or 4 days before 3,4-methylenedioxymethamphetamine (MDMA) (20 mg·kg⁻¹, i.p., two injections separated 3 h) on horizontal locomotor activity measured during the 30 min immediately after the second MDMA injection. Data shown as mean ± SEM, n= 5 − 11. There were no differences between the data obtained from saline controls pretreated 1 or 4 days earlier with cocaine and therefore data were grouped. Different from saline: *P < 0.001; different from MDMA: ^ΔP < 0.001; different from cocaine + MDMA 1 day later: ^δP < 0.001; different from cocaine + saline: ^fP < 0.001. One-way anova followed by Newman–Keuls test.

Figure 5

Effect of repeated cocaine (20 mg·kg⁻¹, i.p. twice daily separated by 8 h for 3 days) finishing 4 days before on 3,4-methylenedioxymethamphetamine (MDMA) (20 mg·kg⁻¹, i.p., two injections separated 3 h)-induced changes in extracellular (A) dopamine (DA), (B) DOPAC and (C) HVA concentration in striatum. MDMA increased extracellular levels of dopamine [F_1,11= 37.8, P < 0.001] and decreased levels of DOPAC [F_1,12= 79.05, P < 0.001] and HVA [F_1,10= 3.61, P < 0.05] compared with saline-treated animals. Pretreatment of the animals with cocaine finishing 4 days before MDMA potentiated this increase in extracellular dopamine levels [F_1,19= 7.53, P < 0.01], but did not modify levels of DOPAC [F_1,18= 0.75, P= 0.36; n.s.] or HVA [F_1,17= 0.20, P= 0.64, n.s.]. Data expressed as % of baseline values. Results represent mean ± SEM, n= 4 − 11; two-way anova. Basal dopamine concentrations were lower in mice pretreated with cocaine compared with saline-treated mice (bar graph). Different from saline-pretreated mice: *P < 0.05 (t-test). Basal DOPAC and HVA levels were similar between groups, for DOPAC (pg·µL⁻¹): 182.0 ± 16.2 versus 185.0 ± 20.0 for saline- and cocaine-pretreated mice, respectively (P= 0.91, n.s., t-test) and for HVA (pg·µL⁻¹): 117.7 ± 13.1 versus 118.8 ± 13.5 for saline- and cocaine-pretreated mice, respectively (P= 0.95, n.s., t-test).