Dopamine release in nucleus accumbens is under tonic inhibition by adenosine A₁ receptors regulated by astrocytic ENT1 and dysregulated by ethanol

Striatal adenosine A1 receptor (A1R) activation can inhibit dopamine release. A1Rs on other striatal neurons are activated by an adenosine tone that is limited by equilibrative nucleoside transporter 1 (ENT1) that is enriched on astrocytes and is ethanol-sensitive. We explored whether dopamine release in nucleus accumbens core is under tonic inhibition by A1Rs, and is regulated by astrocytic ENT1 and ethanol. In ex vivo striatal slices from male and female mice, A1R agonists inhibited dopamine release evoked electrically or optogenetically and detected using fast-scan cyclic voltammetry, most strongly for lower stimulation frequencies and pulse numbers, thereby enhancing the activity-dependent contrast of dopamine release. Conversely, A1R antagonists reduced activity-dependent contrast but enhanced evoked dopamine release levels, even for single optogenetic pulses indicating an underlying tonic inhibition. The ENT1 inhibitor NBTI reduced dopamine release and promoted A1R-mediated inhibition, and conversely, virally-mediated astrocytic overexpression of ENT1 enhanced dopamine release and relieved A1R-mediated inhibition. By imaging the genetically encoded fluorescent adenosine sensor GRAB-Ado, we identified a striatal extracellular adenosine tone that was elevated by the ENT1 inhibitor and sensitive to gliotoxin fluorocitrate. Finally, we identified that ethanol (50 mM) promoted A1R-mediated inhibition of dopamine release, through diminishing adenosine uptake via ENT1. Together, these data reveal that dopamine output dynamics are gated by a striatal adenosine tone, limiting amplitude but promoting contrast, regulated by ENT1, and promoted by ethanol. These data add to the diverse mechanisms through which ethanol modulates striatal dopamine, and to emerging datasets supporting astrocytic transporters as important regulators of striatal function.SIGNIFICANCE STATEMENTDopamine axons in the mammalian striatum are emerging as strategic sites where neuromodulators can powerfully influence dopamine output in health and disease. We found that ambient levels of the neuromodulator adenosine tonically inhibit dopamine release in nucleus accumbens core via adenosine A1 receptors (A1Rs), to a variable level that promotes the contrast in dopamine signals released by different frequencies of activity. We reveal that the equilibrative nucleoside transporter 1 (ENT1) on astrocytes limits this tonic inhibition, and that ethanol promotes it by diminishing adenosine uptake via ENT1. These findings support the hypotheses that A1Rs on dopamine axons inhibit DA release and, furthermore, that astrocytes perform important roles in setting the level of striatal dopamine output, in health and disease.