Aversive behavior induced by optogenetic inactivation of ventral tegmental area dopamine neurons is mediated by dopamine D2 receptors in the nucleus accumbens

Dopamine (DA) transmission from the ventral tegmental area (VTA) is critical for controlling both rewarding and aversive behaviors. The transient silencing of DA neurons is one of the responses to aversive stimuli, but its consequences and neural mechanisms regarding aversive responses and learning have largely remained elusive. Here, we report that optogenetic inactivation of VTA DA neurons promptly down-regulated DA levels and induced up-regulation of the neural activity in the nucleus accumbens (NAc) as evaluated by Fos expression. This optogenetic suppression of DA neuron firing immediately evoked aversive responses to the previously preferred dark room and led to aversive learning toward the optogenetically conditioned place. Importantly, this place aversion was abolished by knockdown of dopamine D2 receptors but not by that of D1 receptors in the NAc. Silencing of DA neurons in the VTA was thus indispensable for inducing aversive responses and learning through dopamine D2 receptors in the NAc.The mesolimbic dopaminergic system not only plays a pivotal role in a wide range of motivation and learning (1–3), but its dysfunction has also been implicated in severe neuropsychiatric disorders as exemplified in Parkinson disease, schizophrenia, and drug addiction. Dopamine (DA) neurons in the ventral tegmental area (VTA) react to rewarding stimuli by phasic firing, and the main function of this firing is theorized to encode “the reward prediction error,” the difference in the value between the predicted reward and the actual reward (4). In contrast to the response to rewarding stimuli, their reactions to aversive stimuli are far from homologous; i.e., some DA neurons are activated in response to aversive stimuli, whereas most others react by transiently suppressing their firings (5–9). In fact, recent studies have revealed that optogenetic activation of GABAergic neurons and resultant inactivation of DA neurons suppress reward consumption and induce an aversive response (10, 11). However, it has largely remained elusive as to which mechanisms in the neural circuits are essential for the acquisition of aversive learning following the inactivation of DA neurons in the VTA and as to how behavioral responses are controlled toward suppressing reward consumption and inducing aversive behaviors.Accumulated evidence has revealed that the motivational and cognitive learning in response to positive and negative stimuli is largely regulated by the neural circuits including the basal ganglia (12), which receive a large amount of the dopaminergic projection from the midbrain. In the striatum, two fundamental neural circuits are constituted by specified medium-sized spiny neurons (MSNs), each expressing a distinct type of DA receptor (13). One circuit is the direct pathway, consisting of the MSNs directly projecting to the output nuclei of the basal ganglia, substantia nigra pars reticulata (SNr), and predominantly expressing dopamine D1 receptors (D1Rs). The other is the indirect pathway, consisting of the MSNs that project indirectly through the globus pallidus to the SNr and primarily express dopamine D2 receptors (D2Rs). DA signals from the midbrain dynamically modulate these two parallel pathways in the opposite manner via D1Rs and D2Rs, and this modulation is supposed to facilitate motivational learning (3, 14). As for the rewarding stimuli, up-regulated DA levels induced by rewarding signals are considered to activate the D1Rs and thus predominantly facilitate the direct pathway in the nucleus accumbens (NAc). On the other hand, the suppression of DA neuron firings in response to aversive stimuli decreases DA levels in the NAc; and this reaction is supposed to specifically promote the signal transmission in the indirect pathway through activated D2Rs.Although studies using the pharmacological strategies and reversible neurotransmission blocking (RNB) method have supported this mechanism of regulation in the NAc (15, 16), it has remained unknown whether the suppression of DA neuron firing is sufficient to promote the activity of the indirect pathway and subsequently induce the avoidance behavior. In this present study, we addressed this issue by selectively inactivating DA neurons in the VTA by optogenetically manipulating membrane-hyperpolarizing Arch protein (17) and explicitly demonstrated that the suppression of DA neurons in the VTA subsequently decreased DA levels in the NAc and induced aversive reaction and learning. Furthermore, we investigated the mechanisms of the regulation of this reaction and disclosed that this aversive reaction was specifically controlled by D2Rs in the NAc.