Inhibition of aldehyde dehydrogenase-2 suppresses cocaine seeking by generating THP, a cocaine use-dependent inhibitor of dopamine synthesis

Abstract

There is no effective treatment for cocaine addiction despite extensive knowledge of the neurobiology of drug addiction. Here we show that a selective aldehyde dehydrogenase-2 (ALDH-2) inhibitor, ALDH2i, suppresses cocaine self-administration in rats and prevents cocaine- or cue-induced reinstatement in a rat model of cocaine relapse-like behavior. We also identify a molecular mechanism by which ALDH-2 inhibition reduces cocaine-seeking behavior: increases in tetrahydropapaveroline (THP) formation due to inhibition of ALDH-2 decrease cocaine-stimulated dopamine production and release in vitro and in vivo. Cocaine increases extracellular dopamine concentration, which activates dopamine D2 autoreceptors to stimulate cAMP-dependent protein kinase A (PKA) and protein kinase C (PKC) in primary ventral tegmental area (VTA) neurons. PKA and PKC phosphorylate and activate tyrosine hydroxylase, further increasing dopamine synthesis in a positive-feedback loop. Monoamine oxidase converts dopamine to 3,4-dihydroxyphenylacetaldehyde (DOPAL), a substrate for ALDH-2. Inhibition of ALDH-2 enables DOPAL to condense with dopamine to form THP in VTA neurons. THP selectively inhibits phosphorylated (activated) tyrosine hydroxylase to reduce dopamine production via negative-feedback signaling. Reducing cocaine- and craving-associated increases in dopamine release seems to account for the effectiveness of ALDH2i in suppressing cocaine-seeking behavior. Selective inhibition of ALDH-2 may have therapeutic potential for treating human cocaine addiction and preventing relapse.

Conflict of interest statement

COMPETING FINANCIAL INTERESTS

The authors declare no competing financial interests.

Figures

Figure 1

ALDH2i reduces intravenous cocaine self-administration, cocaine-primed or cue-induced reinstatement and methamphetamine-induced reinstatement in Sprague Dawley rats. (a) The number of cocaine infusions recorded during the 2-h cocaine self-administration session (n = 7–12, *P < 0.05, **P < 0.01 compared with vehicle (Veh)). (b,c) The number of lever presses recorded during the 2-h cocaine-primed (b) or cue-induced (c) reinstatement session (n = 6–9 and n = 6–11 for b and c, respectively; #P < 0.01 compared with extinction (Ext); **P < 0.01 compared with Veh). (d) The number of lever presses recorded during the 2-h methamphetamine-induced reinstatement session (n = 7, #P < 0.01 compared with Ext; *P < 0.05 compared with Veh).

Figure 2

ALDH2i decreases cocaine-induced dopamine (DA) production and increases THP abundance in PC12 cells. (a–e) Cells were incubated with or without cocaine (Coca, 1 μM) in the presence or absence of ALDH2i, the D1 antagonist SCH 23390 (SCH, 10 μM), the D2 antagonist spiperone (SPIP, 10 μM) or THP for 24 h. Intracellular and extracellular dopamine (a,b,e), total dopamine (c) or THP amounts (d) were determined. *P < 0.05, **P < 0.01 compared with cocaine control. (f) Dose-response analysis of THP and α-methyl-L-tyrosine (αMLT) on the inhibition of total tyrosine hydroxylase (TH) or phosphorylated tyrosine hydroxylase (p-TH).

Figure 3

Cocaine activates PKA and PKC to phosphorylate tyrosine hydroxylase and increase dopamine production in VTA neurons. (a–e) Cells were incubated for 24 h with or without cocaine (1 μM) or nomifensine (NMFS, 10 μM) in the presence or absence of the PKA activator Sp-cAMPS (Sp, 1 mM, 10 min), the PKC activator phorbol 12-myristate 13-acetate (PMA, 100 nM, 10 min), the D1 antagonist SCH23390 (10 μM), the D2 antagonist spiperone (10 μM), the PKA inhibitor Rp-cAMPS (Rp, 20 μM) or the PKC inhibitor GF 109203X (GF, 1 μM). TH phosphorylation at Ser40 p-40; (a–d), at Ser19 (p-19) and Ser31 (p-31; a) or the translocation of PKA and εPKC (e) was detected by western blotting (a,c–e) or by immunostaining (b). Green indicates phosphorylated tyrosine hydroxylase at Ser40 (p-TH), red, total tyrosine hydroxylase and yellow, the merged images. C/P, the ratio of PKA or ePKC in the cytosolic fraction (C) versus the particulate fraction (P). (f,g) Intracellular and extracellular dopamine (f) or total dopamine and THP (g). **P < 0.01 compared with sham or cocaine control.

Figure 4

ALDH2i increases THP production to inhibit tyrosine hydroxylase activity and decrease dopamine production in VTA in cocaine-addicted rats. (a,b) Dopamine and THP amounts measured in the VTA (a) and nucleus accumbens (b) pooled from the cocaine-seeking rats treated with ALDH2i (I, 15 mg per kg body weight, n = 8) or vehicle (V, n = 8) in Figure 1c or from naive rats treated with ALDH2i (15 mg per kg body weight, n = 8) or vehicle (n = 8). The experiment was repeated with similar results. (c) Tyrosine hydroxylase phosphorylation at Ser40 in the VTA, as detected by western blotting. **P < 0.01 compared with naive vehicle control. (d) The number of lever presses during the 2-h cue-induced cocaine reinstatement session (n = 8, #P < 0.01 compared with extinction; **P < 0.01 compared with Veh).