Loss of microglial MCT4 leads to defective synaptic pruning and anxiety-like behavior in mice.

Microglia, the innate immune cells of the central nervous system, actively participate in brain development by supporting neuronal maturation and refining synaptic connections. These cells are emerging as highly metabolically flexible, able to oxidize different energetic substrates to meet their energy demand. Lactate is particularly abundant in the brain, but whether microglia use it as a metabolic fuel has been poorly explored. Here we show that microglia can import lactate, and this is coupled with increased lysosomal acidification. In vitro, loss of the monocarboxylate transporter MCT4 in microglia prevents lactate-induced lysosomal modulation and leads to defective cargo degradation. Microglial depletion of MCT4 in vivo leads to impaired synaptic pruning, associated with increased excitation in hippocampal neurons, enhanced AMPA/GABA ratio, vulnerability to seizures and anxiety-like phenotype. Overall, these findings show that selective disruption of the MCT4 transporter in microglia is sufficient to alter synapse refinement and to induce defects in mouse brain development and adult behavior.

Plasticity of neuronal dynamics in the lateral habenula for cue-punishment associative learning.

The brain's ability to associate threats with external stimuli is vital to execute essential behaviours including avoidance. Disruption of this process contributes instead to the emergence of pathological traits which are common in addiction and depression. However, the mechanisms and neural dynamics at the single-cell resolution underlying the encoding of associative learning remain elusive. Here, employing a Pavlovian discrimination task in mice we investigate how neuronal populations in the lateral habenula (LHb), a subcortical nucleus whose excitation underlies negative affect, encode the association between conditioned stimuli and a punishment (unconditioned stimulus). Large population single-unit recordings in the LHb reveal both excitatory and inhibitory responses to aversive stimuli. Additionally, local optical inhibition prevents the formation of cue discrimination during associative learning, demonstrating a critical role of LHb activity in this process. Accordingly, longitudinal in vivo two-photon imaging tracking LHb calcium neuronal dynamics during conditioning reveals an upward or downward shift of individual neurons' CS-evoked responses. While recordings in acute slices indicate strengthening of synaptic excitation after conditioning, support vector machine algorithms suggest that postsynaptic dynamics to punishment-predictive cues represent behavioral cue discrimination. To examine the presynaptic signaling in LHb participating in learning we monitored neurotransmitter dynamics with genetically-encoded indicators in behaving mice. While glutamate, GABA, and serotonin release in LHb remain stable across associative learning, we observe enhanced acetylcholine signaling developing throughout conditioning. In summary, converging presynaptic and postsynaptic mechanisms in the LHb underlie the transformation of neutral cues in valued signals supporting cue discrimination during learning.

Synaptic inhibition in the lateral habenula shapes reward anticipation.

The lateral habenula (LHb) supports learning processes enabling the prediction of upcoming rewards. While reward-related stimuli decrease the activity of LHb neurons, whether this anchors on synaptic inhibition to guide reward-driven behaviors remains poorly understood. Here, we combine in vivo two-photon calcium imaging with Pavlovian conditioning in mice and report that anticipatory licking emerges along with decreases in cue-evoked calcium signals in individual LHb neurons. In vivo multiunit recordings and pharmacology reveal that the cue-evoked reduction in LHb neuronal firing relies on GABAA-receptor activation. In parallel, we observe a postsynaptic potentiation of GABAA-receptor-mediated inhibition, but not excitation, onto LHb neurons together with the establishment of anticipatory licking. Finally, strengthening or weakening postsynaptic inhibition with optogenetics and GABAA-receptor manipulations enhances or reduces anticipatory licking, respectively. Hence, synaptic inhibition in the LHb shapes reward anticipation.

Mild stress accumulation limits GABAergic synaptic plasticity in the lateral habenula.

Animals can cope with isolated stressful situations without enduring long-term consequences. However, when exposure to stressors becomes recurrent, behavioural symptoms of anxiety and depression can emerge. Yet, the neuronal mechanisms governing responsivity to isolated stressor remain elusive. Here, we investigate synaptic adaptations following mild stress in the lateral habenula (LHb), a structure engaged in aversion encoding and dysfunctional in depression. We describe that neuronal depolarization in the LHb drives long-term depression of inhibitory, but not excitatory, synaptic transmission (GABA LTD). This plasticity requires nitric oxide and presynaptic GABAB receptors, leading to a decrease in probability of GABA release. Mild stressors such as brief social isolation, or exposure to novel environment in the company of littermates, do not alter GABA LTD. In contrast, GABA LTD is absent after mice experience a novel environment in social isolation. Altogether, our results suggest that LHb GABAergic plasticity is sensitive to stress accumulation, which could represent a threshold mechanism for long-term alterations of LHb function.

Septal cholinergic input to CA2 hippocampal region controls social novelty discrimination via nicotinic receptor-mediated disinhibition.

Acetylcholine (ACh), released in the hippocampus from fibers originating in the medial septum/diagonal band of Broca (MSDB) complex, is crucial for learning and memory. The CA2 region of the hippocampus has received increasing attention in the context of social memory. However, the contribution of ACh to this process remains unclear. Here, we show that in mice, ACh controls social memory. Specifically, MSDB cholinergic neurons inhibition impairs social novelty discrimination, meaning the propensity of a mouse to interact with a novel rather than a familiar conspecific. This effect is mimicked by a selective antagonist of nicotinic AChRs delivered in CA2. Ex vivo recordings from hippocampal slices provide insight into the underlying mechanism, as activation of nAChRs by nicotine increases the excitatory drive to CA2 principal cells via disinhibition. In line with this observation, optogenetic activation of cholinergic neurons in MSDB increases the firing of CA2 principal cells in vivo. These results point to nAChRs as essential players in social novelty discrimination by controlling inhibition in the CA2 region.

Stress undermines reward-guided cognitive performance through synaptic depression in the lateral habenula.

Weighing alternatives during reward pursuit is a vital cognitive computation that, when disrupted by stress, yields aspects of neuropsychiatric disorders. To examine the neural mechanisms underlying these phenomena, we employed a behavioral task in which mice were confronted by a reward and its omission (i.e., error). The experience of error outcomes engaged neuronal dynamics within the lateral habenula (LHb), a subcortical structure that supports appetitive behaviors and is susceptible to stress. A high incidence of errors predicted low strength of habenular excitatory synapses. Accordingly, stressful experiences increased error choices while decreasing glutamatergic neurotransmission onto LHb neurons. This synaptic adaptation required a reduction in postsynaptic AMPA receptors (AMPARs), irrespective of the anatomical source of glutamate. Bidirectional control of habenular AMPAR transmission recapitulated and averted stress-driven cognitive deficits. Thus, a subcortical synaptic mechanism vulnerable to stress underlies behavioral efficiency during cognitive performance.

Morphine withdrawal recruits lateral habenula cytokine signaling to reduce synaptic excitation and sociability.

The lateral habenula encodes aversive stimuli contributing to negative emotional states during drug withdrawal. Here we report that morphine withdrawal in mice leads to microglia adaptations and diminishes glutamatergic transmission onto raphe-projecting lateral habenula neurons. Chemogenetic inhibition of this circuit promotes morphine withdrawal-like social deficits. Morphine withdrawal-driven synaptic plasticity and reduced sociability require tumor necrosis factor-α (TNF-α) release and neuronal TNF receptor 1 activation. Hence, habenular cytokines control synaptic and behavioral adaptations during drug withdrawal.

Punishment-Predictive Cues Guide Avoidance through Potentiation of Hypothalamus-to-Habenula Synapses.

Throughout life, individuals learn to predict a punishment via its association with sensory stimuli. This process ultimately prompts goal-directed actions to prevent the danger, a behavior defined as avoidance. Neurons in the lateral habenula (LHb) respond to aversive events as well as to environmental cues predicting them, supporting LHb contribution to cue-punishment association. However, whether synaptic adaptations at discrete habenular circuits underlie such associative learning to instruct avoidance remains elusive. Here, we find that, in mice, contingent association of an auditory cue (tone) with a punishment (foot shock) progressively causes cue-driven LHb neuronal excitation during avoidance learning. This process is concomitant with the strengthening of LHb AMPA receptor-mediated neurotransmission. Such a phenomenon occludes long-term potentiation and occurs specifically at hypothalamus-to-habenula synapses. Silencing hypothalamic-to-habenulainputs or optically inactivating postsynaptic AMPA receptors within the LHb disrupts avoidance learning. Altogether, synaptic strengthening at a discrete habenular circuit transforms neutral stimuli into salient punishment-predictive cues to guide avoidance.

Motor thalamus supports striatum-driven reinforcement.

Reinforcement has long been thought to require striatal synaptic plasticity. Indeed, direct striatal manipulations such as self-stimulation of direct-pathway projection neurons (dMSNs) are sufficient to induce reinforcement within minutes. However, it's unclear what role, if any, is played by downstream circuitry. Here, we used dMSN self-stimulation in mice as a model for striatum-driven reinforcement and mapped the underlying circuitry across multiple basal ganglia nuclei and output targets. We found that mimicking the effects of dMSN activation on downstream circuitry, through optogenetic suppression of basal ganglia output nucleus substantia nigra reticulata (SNr) or activation of SNr targets in the brainstem or thalamus, was also sufficient to drive rapid reinforcement. Remarkably, silencing motor thalamus-but not other selected targets of SNr-was the only manipulation that reduced dMSN-driven reinforcement. Together, these results point to an unexpected role for basal ganglia output to motor thalamus in striatum-driven reinforcement.

Pathway-Specific Remodeling of Thalamostriatal Synapses in Parkinsonian Mice.

Movement suppression in Parkinson's disease (PD) is thought to arise from increased efficacy of the indirect pathway basal ganglia circuit, relative to the direct pathway. However, the underlying pathophysiological mechanisms remain elusive. To examine whether changes in the strength of synaptic inputs to these circuits contribute to this imbalance, we obtained paired whole-cell recordings from striatal direct- and indirect-pathway medium spiny neurons (dMSNs and iMSNs) and optically stimulated inputs from sensorimotor cortex or intralaminar thalamus in brain slices from control and dopamine-depleted mice. We found that dopamine depletion selectively decreased synaptic strength at thalamic inputs to dMSNs, suggesting that thalamus drives asymmetric activation of basal ganglia circuitry underlying parkinsonian motor impairments. Consistent with this hypothesis, in vivo chemogenetic and optogenetic inhibition of thalamostriatal terminals reversed motor deficits in dopamine-depleted mice. These results implicate thalamostriatal projections in the pathophysiology of PD and support interventions targeting thalamus as a potential therapeutic strategy.

Cell-Type-Specific Control of Brainstem Locomotor Circuits by Basal Ganglia.

The basal ganglia (BG) are critical for adaptive motor control, but the circuit principles underlying their pathway-specific modulation of target regions are not well understood. Here, we dissect the mechanisms underlying BG direct and indirect pathway-mediated control of the mesencephalic locomotor region (MLR), a brainstem target of BG that is critical for locomotion. We optogenetically dissect the locomotor function of the three neurochemically distinct cell types within the MLR: glutamatergic, GABAergic, and cholinergic neurons. We find that the glutamatergic subpopulation encodes locomotor state and speed, is necessary and sufficient for locomotion, and is selectively innervated by BG. We further show activation and suppression, respectively, of MLR glutamatergic neurons by direct and indirect pathways, which is required for bidirectional control of locomotion by BG circuits. These findings provide a fundamental understanding of how BG can initiate or suppress a motor program through cell-type-specific regulation of neurons linked to specific actions.

Firing modes of dopamine neurons drive bidirectional GIRK channel plasticity.

G-protein-coupled inwardly rectifying potassium (GIRK) channels contribute to the resting membrane potential of many neurons, including dopamine (DA) neurons in the ventral tegmental area (VTA). VTA DA neurons are bistable, firing in two modes: one characterized by bursts of action potentials, the other by tonic firing at a lower frequency. Here we provide evidence that these firing modes drive bidirectional plasticity of GIRK channel-mediated currents. In acute midbrain slices of mice, we observed that in vitro burst activation of VTA DA neurons potentiated GIRK currents whereas tonic firing depressed these currents. This plasticity was not specific to the metabotropic receptor activating the GIRK channels, as direct activation of GIRK channels by nonhydrolyzable GTP also potentiated the currents. The plasticity of GIRK currents required NMDA receptor and CaMKII activation, and involved protein trafficking through specific PDZ domains of GIRK2c and GIRK3 subunit isoforms. Prolonged tonic firing may thus enhance the probability to switch into burst-firing mode, which then potentiates GIRK currents and favors the return to baseline. In conclusion, activity-dependent GIRK channel plasticity may represent a slow destabilization process favoring the switch between the two firing modes of VTA DA neurons.

Methamphetamine-evoked depression of GABA(B) receptor signaling in GABA neurons of the VTA.

Psychostimulants induce neuroadaptations in excitatory and fast inhibitory transmission in the ventral tegmental area (VTA). Mechanisms underlying drug-evoked synaptic plasticity of slow inhibitory transmission mediated by GABA(B) receptors and G protein-gated inwardly rectifying potassium (GIRK/Kir(3)) channels, however, are poorly understood. Here, we show that 1 day after methamphetamine (METH) or cocaine exposure both synaptically evoked and baclofen-activated GABA(B)R-GIRK currents were significantly depressed in VTA GABA neurons and remained depressed for 7 days. Presynaptic inhibition mediated by GABA(B)Rs on GABA terminals was also weakened. Quantitative immunoelectron microscopy revealed internalization of GABA(B1) and GIRK2, which occurred coincident with dephosphorylation of serine 783 (S783) in GABA(B2), a site implicated in regulating GABA(B)R surface expression. Inhibition of protein phosphatases recovered GABA(B)R-GIRK currents in VTA GABA neurons of METH-injected mice. This psychostimulant-evoked impairment in GABA(B)R signaling removes an intrinsic brake on GABA neuron spiking, which may augment GABA transmission in the mesocorticolimbic system.

Is there a way to curb benzodiazepine addiction?

Benzodiazepines are widely prescribed drugs used to treat anxiety and insomnia, induce muscle relaxation, control epileptic seizures, promote anaesthesia or produce amnesia. Benzodiazepines are also abused for recreational purposes and the number of benzodiazepine abusers is unfortunately increasing. Within weeks of chronic use, tolerance to the pharmacological effects can develop and withdrawal becomes apparent once the drug is no longer available, which are both conditions indicative of benzodiazepine dependence. Diagnosis of addiction (i.e. compulsive use despite negative consequences) may follow in vulnerable individuals. Here, we review the historical and current use of benzodiazepines from their original synthesis, discovery and commercialisation to the recent identification of the molecular mechanism by which benzodiazepines induce addiction. These results have identified the mechanisms underlying the activation of midbrain dopamine neurons by benzodiazepines, and how these drugs can hijack the mesocorticolimbic reward system. Such knowledge calls for future developments of new receptor subtype specific benzodiazepines with a reduced addiction liability.

Morphine- and CaMKII-dependent enhancement of GIRK channel signaling in hippocampal neurons.

G-protein-gated inwardly rectifying potassium (GIRK) channels, which help control neuronal excitability, are important for the response to drugs of abuse. Here, we describe a novel pathway for morphine-dependent enhancement of GIRK channel signaling in hippocampal neurons. Morphine treatment for ∼20 h increased the colocalization of GIRK2 with PSD95, a dendritic spine marker. Western blot analysis and quantitative immunoelectron microscopy revealed an increase in GIRK2 protein and targeting to dendritic spines. In vivo administration of morphine also produced an upregulation of GIRK2 protein in the hippocampus. The mechanism engaged by morphine required elevated intracellular Ca(2+) and was insensitive to pertussis toxin, implicating opioid receptors that may couple to Gq G-proteins. Met-enkephalin, but not the µ-selective (DAMGO) and δ-selective (DPDPE) opioid receptor agonists, mimicked the effect of morphine, suggesting involvement of a heterodimeric opioid receptor complex. Peptide (KN-93) inhibition of CaMKII prevented the morphine-dependent change in GIRK localization, whereas expression of a constitutively activated form of CaMKII mimicked the effects of morphine. Coincident with an increase in GIRK2 surface expression, functional analyses revealed that morphine treatment increased the size of serotonin-activated GIRK currents and Ba(2+)-sensitive basal K(+) currents in neurons. These results demonstrate plasticity in neuronal GIRK signaling that may contribute to the abusive effects of morphine.