Neurotoxic astrocytes express the d-serine synthesizing enzyme, serine racemase, in Alzheimer's disease.

Although ß-amyloid plaques are a well-recognized hallmark of Alzheimer's disease (AD) neuropathology, no drugs reducing amyloid burden have shown efficacy in clinical trials, suggesting that once AD symptoms emerge, disease progression becomes independent of Aß production. Reactive astrocytes are another neuropathological feature of AD, where there is an emergence of neurotoxic (A1) reactive astrocytes. We find that serine racemase (SR), the neuronal enzyme that produces the N-methyl-d-aspartate receptor (NMDAR) co-agonist d-serine, is robustly expressed in A1-reactive neurotoxic astrocytes in the hippocampus and entorhinal cortex of AD subjects and an AD rat model. Furthermore, we observe intracellular signaling changes consistent with increased extra-synaptic NMDAR activation, excitotoxicity and decreased neuronal survival. Thus, reducing neurotoxic d-serine release from A1 inflammatory astrocytes could have therapeutic benefit for mild to advanced AD, when anti-amyloid strategies are ineffective.

Deletion of Dtnbp1 in mice impairs threat memory consolidation and is associated with enhanced inhibitory drive in the amygdala.

Schizophrenia is a severe and highly heritable disorder. Dystrobrevin-binding protein 1 (DTNBP1), also known as dysbindin-1, has been implicated in the pathophysiology of schizophrenia. Specifically, dysbindin-1 mRNA and protein expression are decreased in the brains of subjects with this disorder. Mice lacking dysbinidn-1 also display behavioral phenotypes similar to those observed in schizophrenic patients. However, it remains unknown whether deletion of dysbindin-1 impacts functions of the amygdala, a brain region that is critical for emotional processing, which is disrupted in patients with schizophrenia. Here, we show that dysbindin-1 is expressed in both excitatory and inhibitory neurons of the basolateral amygdala (BLA). Deletion of dysbindin-1 in male mice (Dys-/-) impaired cued and context-dependent threat memory, without changes in measures of anxiety. The behavioral deficits observed in Dys-/- mice were associated with perturbations in the BLA, including the enhancement of GABAergic inhibition of pyramidal neurons, increased numbers of parvalbumin interneurons, and morphological abnormalities of dendritic spines on pyramidal neurons. Our findings highlight an important role for dysbindin-1 in the regulation of amygdalar function and indicate that enhanced inhibition of BLA pyramidal neuron activity may contribute to the weakened threat memory expression observed in Dys-/- mice.

Bidirectional and long-lasting control of alcohol-seeking behavior by corticostriatal LTP and LTD.

Addiction is proposed to arise from alterations in synaptic strength via mechanisms of long-term potentiation (LTP) and depression (LTD). However, the causality between these synaptic processes and addictive behaviors is difficult to demonstrate. Here we report that LTP and LTD induction altered operant alcohol self-administration, a motivated drug-seeking behavior. We first induced LTP by pairing presynaptic glutamatergic stimulation with optogenetic postsynaptic depolarization in the dorsomedial striatum, a brain region known to control goal-directed behavior. Blockade of this LTP by NMDA-receptor inhibition unmasked an endocannabinoid-dependent LTD. In vivo application of the LTP-inducing protocol caused a long-lasting increase in alcohol-seeking behavior, while the LTD protocol decreased this behavior. We further identified that optogenetic LTP and LTD induction at cortical inputs onto striatal dopamine D1 receptor-expressing neurons controlled these behavioral changes. Our results demonstrate a causal link between synaptic plasticity and alcohol-seeking behavior and suggest that modulation of this plasticity may inspire a therapeutic strategy for addiction.

Dopamine D1 or D2 receptor-expressing neurons in the central nervous system.

Dopamine signals mainly through D1 receptors (D1Rs) and D2 receptors (D2Rs); D1R-expressing or D2R-expressing neurons contribute to distinct reward and addictive behaviors. Traditionally, transgenic mice expressing green fluorescent protein (GFP) under D1R or D2R promoters are used for fluorescent verification in electrophysiology studies, whereas Cre mice are employed for behavioral research. However, it is unknown whether the same neuronal populations are targeted in GFP and Cre mice. Additionally, while D1Rs and D2Rs are known to be expressed in different striatal neurons, their expression patterns outside the striatum remain unclear. The present study addressed these two questions by using several transgenic mouse lines expressing fluorescent proteins (GFP or tdTomato) or Cre under the control of D1R or D2R promoters. We found a high degree of overlap between GFP-positive and Cre-positive neurons in the striatum and hippocampus. Additionally, we discovered that D1Rs and D2Rs were highly segregated in the orbitofrontal cortex, prefrontal cortex, dorsal and ventral hippocampus, and amygdala: ~4-34 percent of neurons co-expressed these receptors. Importantly, slice electrophysiological studies demonstrated that D1R-positive and D1R-negative hippocampal neurons were functionally distinct in a mouse line generated by crossing Drd1a-Cre mice with a Cre reporter Ai14 line. Lastly, we discovered that chronic alcohol intake differentially altered D1R-positive and D2R-positive neuron excitability in the ventral CA1. These data suggest that GFP and Cre mice target the same populations of striatal neurons, D1R-expressing or D2R-expressing neurons are highly segregated outside the striatum, and these neurons in the ventral hippocampal may exert distinct roles in alcohol addiction.

Serine Racemase and D-serine in the Amygdala Are Dynamically Involved in Fear Learning.

BACKGROUND: The amygdala is a central component of the neural circuitry that underlies fear learning. N-methyl-D-aspartate receptor-dependent plasticity in the amygdala is required for pavlovian fear conditioning and extinction. N-methyl-D-aspartate receptor activation requires the binding of a coagonist, D-serine, which is synthesized from L-serine by the neuronal enzyme serine racemase (SR). However, little is known about SR and D-serine function in the amygdala. METHODS: We used immunohistochemical methods to characterize the cellular localization of SR and D-serine in the mouse and human amygdala. Using biochemical and molecular techniques, we determined whether trace fear conditioning and extinction engages the SR/D-serine system in the brain. D-serine was administered systemically to mice to evaluate its effect on fear extinction. Finally, we investigated whether the functional single nucleotide polymorphism rs4523957, which is an expression quantitative trait locus of the human serine racemase (SRR) gene, was associated with fear-related phenotypes in a highly traumatized human cohort. RESULTS: We demonstrate that approximately half of the neurons in the amygdala express SR, including both excitatory and inhibitory neurons. We find that the acquisition and extinction of fear memory engages the SR/D-serine system in the mouse amygdala and that D-serine administration facilitates fear extinction. We also demonstrate that the SRR single nucleotide polymorphism, rs4523957, is associated with posttraumatic stress disorder in humans, consistent with the facilitatory effect of D-serine on fear extinction. CONCLUSIONS: These new findings have important implications for understanding D-serine-mediated N-methyl-D-aspartate receptor plasticity in the amygdala and how this system could contribute to disorders with maladaptive fear circuitry.

Stroke triggers nigrostriatal plasticity and increases alcohol consumption in rats.

Excessive alcohol consumption is a known risk factor for stroke, but the effect of stroke on alcohol intake is unknown. The dorsomedial striatum (DMS) and midbrain areas of the nigrostriatal circuit are critically associated to stroke and alcohol addiction. Here we sought to explore the influence of stroke on alcohol consumption and to uncover the underlying nigrostriatal mechanism. Rats were trained to consume alcohol using a two-bottle choice or operant self-administration procedure. Retrograde beads were infused into the DMS or midbrain to label specific neuronal types, and ischemic stroke was induced in the dorsolateral striatum (DLS). Slice electrophysiology was employed to measure excitability and synaptic transmission in DMS and midbrain neurons. We found that ischemic stroke-induced DLS infarction produced significant increases in alcohol preference, operant self-administration, and relapse. These increases were accompanied by enhanced excitability of DMS and midbrain neurons. In addition, glutamatergic inputs onto DMS D1-neurons was potentiated, whereas GABAergic inputs onto DMS-projecting midbrain dopaminergic neurons was suppressed. Importantly, systemic inhibition of dopamine D1 receptors attenuated the stroke-induced increase in operant alcohol self-administration. Our results suggest that the stroke-induced DLS infarction evoked abnormal plasticity in nigrostriatal dopaminergic neurons and DMS D1-neurons, contributing to increased post-stroke alcohol-seeking and relapse.

Distinct Synaptic Strengthening of the Striatal Direct and Indirect Pathways Drives Alcohol Consumption.

BACKGROUND: Repeated exposure to addictive drugs or alcohol triggers glutamatergic and gamma-aminobutyric acidergic (GABAergic) plasticity in many neuronal populations. The dorsomedial striatum (DMS), a brain region critically involved in addiction, contains medium spiny neurons (MSNs) expressing dopamine D1 or D2 receptors, which form direct and indirect pathways, respectively. It is unclear how alcohol-evoked plasticity in the DMS contributes to alcohol consumption in a cell type-specific manner. METHODS: Mice were trained to consume alcohol using an intermittent-access two-bottle-choice drinking procedure. Slice electrophysiology was used to measure glutamatergic and GABAergic strength in DMS D1- and D2-MSNs of alcohol-drinking mice and control mice. In vivo chemogenetic and pharmacologic approaches were employed to manipulate MSN activity, and their consequences on alcohol consumption were measured. RESULTS: Repeated cycles of alcohol consumption and withdrawal in mice strengthened glutamatergic transmission in D1-MSNs and GABAergic transmission in D2-MSNs. In vivo chemogenetic excitation of D1-MSNs, mimicking glutamatergic strengthening, promoted alcohol consumption; the same effect was induced by D2-MSN inhibition, mimicking GABAergic strengthening. Importantly, suppression of GABAergic transmission via D2 receptor-glycogen synthase kinase-3ß signaling dramatically reduced excessive alcohol consumption, as did selective inhibition of D1-MSNs or excitation of D2-MSNs. CONCLUSIONS: Our results suggest that repeated cycles of excessive alcohol intake and withdrawal potentiate glutamatergic strength exclusively in D1-MSNs and GABAergic strength specifically in D2-MSNs of the DMS, which concurrently contribute to alcohol consumption. These results provide insight into the synaptic and cell type-specific mechanisms underlying alcohol addiction and identify targets for the development of new therapeutic approaches to alcohol abuse.

A new role for AMP-activated protein kinase in the circadian regulation of L-type voltage-gated calcium channels in late-stage embryonic retinal photoreceptors.

AMP-activated protein kinase (AMPK) is a cellular energy sensor, which is activated when the intracellular ATP production decreases. The activities of AMPK display circadian rhythms in various organs and tissues, indicating that AMPK is involved in the circadian regulation of cellular metabolism. In vertebrate retina, the circadian clocks regulate many aspects of retinal function and physiology, including light/dark adaption, but whether and how AMPK was involved in the retinal circadian rhythm was not known. We hypothesized that the activation of AMPK (measured as phosphorylated AMPK) in the retina was under circadian control, and AMPK might interact with other intracellular signaling molecules to regulate photoreceptor physiology. We combined ATP assays, western blots, immunostaining, patch-clamp recordings, and pharmacological treatments to decipher the role of AMPK in the circadian regulation of photoreceptor physiology. We found that the overall retinal ATP content displayed a diurnal rhythm that peaked at early night, which was nearly anti-phase to the diurnal and circadian rhythms of AMPK phosphorylation. AMPK was also involved in the circadian phase-dependent regulation of photoreceptor L-type voltage-gated calcium channels (L-VGCCs), the ion channel essential for sustained neurotransmitter release. The activation of AMPK dampened the L-VGCC currents at night with a corresponding decrease in protein expression of the L-VGCCα1 pore-forming subunit, while inhibition of AMPK increased the L-VGCC current during the day. AMPK appeared to be upstream of extracellular-signal-regulated kinase and mammalian/mechanistic target of rapamycin complex 1 (mTORC1) but downstream of adenylyl cyclase in regulating the circadian rhythm of L-VGCCs. Hence, as a cellular energy sensor, AMPK integrates into the cell signaling network to regulate the circadian rhythm of photoreceptor physiology. We found that in chicken embryonic retina, the activation of AMP-activated protein kinase (AMPK) is under circadian control and anti-phase to the retinal ATP rhythm. While ATP content is higher at night, phosphorylated AMPK (pAMPK) is higher during the day. AMPK appears to be upstream of extracellular signal-regulated kinase (ERK), protein kinase B (AKT), and mammalian target of rapamycin complex 1 (mTORC1) but downstream of adenylyl cyclase in regulating the circadian rhythm of L-VGCCs. Therefore, as a cellular energy sensor, AMPK integrates into the cell signaling network to regulate the circadian rhythm of photoreceptor physiology.

Circadian phase-dependent effect of nitric oxide on L-type voltage-gated calcium channels in avian cone photoreceptors.

Nitric oxide (NO) plays an important role in phase-shifting of circadian neuronal activities in the suprachiasmatic nucleus and circadian behavior activity rhythms. In the retina, NO production is increased in a light-dependent manner. While endogenous circadian oscillators in retinal photoreceptors regulate their physiological states, it is not clear whether NO also participates in the circadian regulation of photoreceptors. In this study, we demonstrate that NO is involved in the circadian phase-dependent regulation of L-type voltage-gated calcium channels (L-VGCCs). In chick cone photoreceptors, the L-VGCCα1 subunit expression and the maximal L-VGCC currents are higher at night, and both Ras-mitogen-activated protein kinase (MAPK)-extracellular signal-regulated kinase (Erk) and Ras-phosphatidylinositol 3 kinase (PI3K)-protein kinase B (Akt) are part of the circadian output pathways regulating L-VGCCs. The NO-cGMP-protein kinase G (PKG) pathway decreases L-VGCCα1 subunit expression and L-VGCC currents at night, but not during the day, and exogenous NO donor or cGMP decreases the phosphorylation of Erk and Akt at night. The protein expression of neural NO synthase (nNOS) is also under circadian control, with both nNOS and NO production being higher during the day. Taken together, NO/cGMP/PKG signaling is involved as part of the circadian output pathway to regulate L-VGCCs in cone photoreceptors. In cone photoreceptors, the protein expression of neural nitric oxide synthase (nNOS) and NO production are under circadian control. NO-cGMP-protein kinase G (PKG) signaling serves in the circadian output pathway to regulate the circadian rhythms of L-type voltage-gated calcium channels (L-VGCCs) in part through regulating the phosphorylation states of extracellular-signal-regulated kinase (Erk) and protein kinase B (Akt).