Impaired spatial memory in adult vitamin D deficient BALB/c mice is associated with reductions in spine density, nitric oxide, and neural nitric oxide synthase in the hippocampus

Vitamin D deficiency is prevalent in adults and is associated with cognitive impairment. However, the mechanism by which adult vitamin D (AVD) deficiency affects cognitive function remains unclear. We examined spatial memory impairment in AVD-deficient BALB/c mice and its underlying mechanism by measuring spine density, long term potentiation (LTP), nitric oxide (NO), neuronal nitric oxide synthase (nNOS) and endothelial NOS (eNOS) in the hippocampus. Adult male BALB/c mice were fed a control or vitamin D deficient diet for 20 weeks. Spatial memory performance was measured using an active place avoidance (APA) task, where AVD-deficient mice had reduced latency entering the shock zone compared to controls. We characterised hippocampal spine morphology in the CA1 and dentate gyrus (DG) and made electrophysiological recordings in the hippocampus of behaviourally naive mice to measure LTP. We next measured NO, as well as glutathione, lipid peroxidation and oxidation of protein products and quantified hippocampal immunoreactivity for nNOS and eNOS. Spine morphology analysis revealed a significant reduction in the number of mushroom spines in the CA1 dendrites but not in the DG. There was no effect of diet on LTP. However, hippocampal NO levels were depleted whereas other oxidation markers were unaltered by AVD deficiency. We also showed a reduced nNOS, but not eNOS, immunoreactivity. Finally, vitamin D supplementation for 10 weeks to AVD-deficient mice restored nNOS immunoreactivity to that seen in in control mice. Our results suggest that lower levels of NO, reduced nNOS immunostaining contribute to hippocampal-dependent spatial learning deficits in AVD-deficient mice.

Dopamine D2L Receptor Deficiency Alters Neuronal Excitability and Spine Formation in Mouse Striatum

The striatum contains several types of neurons including medium spiny projection neurons (MSNs), cholinergic interneurons (ChIs), and fast-spiking interneurons (FSIs). Modulating the activity of these neurons by the dopamine D2 receptor (D2R) can greatly impact motor control and movement disorders. D2R exists in two isoforms: D2L and D2S. Here, we assessed whether alterations in the D2L and D2S expression levels affect neuronal excitability and synaptic function in striatal neurons. We observed that quinpirole inhibited the firing rate of all three types of striatal neurons in wild-type (WT) mice. However, in D2L knockout (KO) mice, quinpirole enhanced the excitability of ChIs, lost influence on spike firing of MSNs, and remained inhibitory effect on spike firing of FSIs. Additionally, we showed mIPSC frequency (but not mIPSC amplitude) was reduced in ChIs from D2L KO mice compared with WT mice, suggesting spontaneous GABA release is reduced at GABAergic terminals onto ChIs in D2L KO mice. Furthermore, we found D2L deficiency resulted in reduced dendritic spine density in ChIs, suggesting D2L activation plays a role in the formation/maintenance of dendritic spines of ChIs. These findings suggest new molecular and cellular mechanisms for causing ChIs abnormality seen in Parkinson’s disease or drug-induced dyskinesias.

Repeated intermittent administration of (R)-ketamine during juvenile and adolescent stages prevents schizophrenia-relevant phenotypes in adult offspring after maternal immune activation: a role of TrkB signaling

Maternal immune activation (MIA) plays a role in the etiology of schizophrenia. MIA by prenatal exposure of polyinosinic:polycytidylic acid [poly(I:C)] in rodents caused behavioral and neurobiological changes relevant to schizophrenia in adult offspring. We investigated whether the novel antidepressant (R)-ketamine could prevent the development of psychosis-like phenotypes in adult offspring after MIA. We examined the effects of (R)-ketamine (10 mg/kg/day, twice weekly for 4 weeks) during juvenile and adolescent stages (P28–P56) on the development of cognitive deficits, loss of parvalbumin (PV)-immunoreactivity in the medial prefrontal cortex (mPFC), and decreased dendritic spine density in the mPFC and hippocampus from adult offspring after prenatal poly(I:C) exposure. Furthermore, we examined the role of TrkB in the prophylactic effects of (R)-ketamine. Repeated intermittent administration of (R)-ketamine during juvenile and adolescent stages significantly blocked the development of cognitive deficits, reduced PV-immunoreactivity in the prelimbic (PrL) of mPFC, and decreased dendritic spine density in the PrL of mPFC, CA3 and dentate gyrus of the hippocampus from adult offspring after prenatal poly(I:C) exposure. Furthermore, pretreatment with ANA-12 (TrkB antagonist: twice weekly for 4 weeks) significantly blocked the beneficial effects of (R)-ketamine on cognitive deficits of adult offspring after prenatal poly(I:C) exposure. These data suggest that repeated intermittent administration of (R)-ketamine during juvenile and adolescent stages could prevent the development of psychosis in adult offspring after MIA. Therefore, (R)-ketamine would be a potential prophylactic drug for young subjects with high-risk for psychosis.

Regulation of Life Extension Factor Klotho on Depressive-like Behaviors via Modulation of GluN2B Function in the Nucleus Accumbens

Klotho is a life extension factor that has an ability to regulate the function of GluN2B-containing N-methyl-D-aspartate receptors (NMDARs), whose dysfunction in the nucleus accumbens (NAc) underlies critical aspects of the pathophysiology of major depression. Here we study the functional relevance of klotho in the pathogenesis of depression. A chronic social defeat stress paradigm, where mice are either categorized as susceptible or unsusceptible group based on their performance in a social interaction test, was used in this study. We found that the expression of klotho was largely decreased in the NAc of susceptible mice when compared to control or unsusceptible group. Genetic knockdown of klotho in the NAc induced depressive-like behaviors in naive mice, while overexpression of klotho produced an antidepressive effect in normal mice and ameliorated the depressive-like behaviors in susceptible mice. Molecularly, knockdown of klotho in the NAc resulted in selective decreases of total and synaptic GluN2B expression that were identical to susceptible mice. Elevation of klotho in the NAc reversed the reductions of GluN2B expressions, as well as altered synaptic transmission and spine density in the NAc of susceptible mice. Furthermore, blockade of GluN2B with a specific antagonist abolished the beneficial effects of klotho elevation in susceptible mice. Collectively, we demonstrated that klotho in the NAc modulates depressive-like behaviors by regulating the function of GluN2B-containing NMDARs. These results reveal a novel role for klotho in the pathogenesis of depression, opening new insights into the molecular basis of major depression.

Acute Ethanol Exposure during Synaptogenesis Rapidly Alters Medium Spiny Neuron Morphology and Synaptic Protein Expression in the Dorsal Striatum

Fetal alcohol spectrum disorders are caused by the disruption of normal brain development in utero. The severity and range of symptoms is dictated by both the dosage and timing of ethanol administration, and the resulting developmental processes that are impacted. In order to investigate the effects of an acute, high-dose intoxication event on the development of medium spiny neurons (MSNs) in the striatum, mice were injected with ethanol on P6, and neuronal morphology was assessed after 24 h, or at 1 month or 5 months of age. Data indicate an immediate increase in MSN dendritic length and branching, a rapid decrease in spine number, and increased levels of the synaptic protein PSD-95 as a consequence of this neonatal exposure to ethanol, but these differences do not persist into adulthood. These results demonstrate a rapid neuronal response to ethanol exposure and characterize the dynamic nature of neuronal architecture in the MSNs. Although differences in neuronal branching and spine density induced by ethanol resolve with time, early changes in the caudate/putamen region have a potential impact on the execution of complex motor skills, as well as aspects of long-term learning and addictive behavior.

NRF2 Activation Ameliorates Oxidative Stress and Improves Mitochondrial Function and Synaptic Plasticity, and in A53T α-Synuclein Hippocampal Neurons

In Parkinson’s disease (PD), brain oxidative stress and mitochondrial dysfunction contribute to neuronal loss as well as motor and cognitive deficits. The transcription factor NRF2 has emerged as a promising therapeutic target in PD because it sits at the intersection of antioxidant and mitochondrial pathways. Here, we investigate the effects of modulating NRF2 activity in neurons isolated from a A53T α-synuclein (A53TSyn) mouse model of synucleinopathy. Embryonic hippocampal neurons were isolated from A53TSyn mice and their wild type (WT) littermates. Neurons were treated with either the NRF2 activator dimethyl fumarate (DMF) or the NRF2 inhibitor ML385. Reactive oxygen species (ROS), dendritic arborization and dendritic spine density were quantified. Mitochondrial bioenergetics were also profiled in these neurons. A53TSyn neurons had increased ROS and reduced basal and maximal mitochondrial respiration relative to WT neurons. A53TSyn neurons also displayed decreased dendritic arborization and reduced spine density. Treatment with DMF reduced ROS levels and improved both mitochondrial function and arborization, while inhibition of NRF2 with ML385 exacerbated these endpoints. Modulation of NRF2 activity had a significant effect on mitochondrial function, oxidative stress, and synaptic plasticity in A53TSyn neurons. These data suggest that NRF2 may be a viable target for therapeutic interventions in PD.

Dopamine D4 Receptor Is a Regulator of Morphine-Induced Plasticity in the Rat Dorsal Striatum

Long-term exposition to morphine elicits structural and synaptic plasticity in reward-related regions of the brain, playing a critical role in addiction. However, morphine-induced neuroadaptations in the dorsal striatum have been poorly studied despite its key function in drug-related habit learning. Here, we show that prolonged treatment with morphine triggered the retraction of the dendritic arbor and the loss of dendritic spines in the dorsal striatal projection neurons (MSNs). In an attempt to extend previous findings, we also explored whether the dopamine D4 receptor (D4R) could modulate striatal morphine-induced plasticity. The combined treatment of morphine with the D4R agonist PD168,077 produced an expansion of the MSNs dendritic arbors and restored dendritic spine density. At the electrophysiological level, PD168,077 in combination with morphine altered the electrical properties of the MSNs and decreased their excitability. Finally, results from the sustantia nigra showed that PD168,077 counteracted morphine-induced upregulation of μ opioid receptors (MOR) in striatonigral projections and downregulation of G protein-gated inward rectifier K+ channels (GIRK1 and GIRK2) in dopaminergic cells. The present results highlight the key function of D4R modulating morphine-induced plasticity in the dorsal striatum. Thus, D4R could represent a valuable pharmacological target for the safety use of morphine in pain management.

Effects of Aging on the Structure and Expression of NMDA Receptors of Somatostatin Expressing Neurons in the Mouse Hippocampus

Changes in the physiology, neurochemistry and structure of neurons, particularly of their dendritic spines, are thought to be crucial players in age-related cognitive decline. One of the most studied brain structures affected by aging is the hippocampus, known to be involved in different essential cognitive processes. While the aging-associated quantitative changes in dendritic spines of hippocampal pyramidal cells have already been studied, the relationship between aging and the structural dynamics of hippocampal interneurons remains relatively unknown. Spines are not a frequent feature in cortical inhibitory neurons, but these postsynaptic structures are abundant in a subpopulation of somatostatin expressing interneurons, particularly in oriens-lacunosum moleculare (O-LM) cells in the hippocampal CA1. Previous studies from our laboratory have shown that the spines of these interneurons are highly plastic and influenced by NMDA receptor manipulation. Thus, in the present study, we have investigated the impact of aging on this interneuronal subpopulation. The analyses were performed in 3−, 9−, and 16-month-old GIN mice, a strain in which somatostatin positive interneurons express GFP. We studied the changes in the density of dendritic spines, en passant boutons, and the expression of NMDA receptors (GluN1 and GluN2B) using confocal microscopy and image analysis. We observed a significant decrease in dendritic spine density in 9-month-old animals when compared with 3-month-old animals. We also observed a decrease in the expression of the GluN2B subunit in O-LM cells, but not of that of GluN1, during aging. These results will constitute the basis for more advanced studies of the structure and connectivity of interneurons during aging and their contribution to cognitive decline.