Noncanonical Metabotropic Glutamate Receptor 5 Signaling in Alzheimer's Disease.

Metabotropic glutamate receptor 5 (mGluR5) is ubiquitously expressed in brain regions responsible for memory and learning. It plays a key role in modulating rapid changes in synaptic transmission and plasticity. mGluR5 supports long-term changes in synaptic strength by regulating the transcription and translation of essential synaptic proteins. ß-Amyloid 42 (Aß42) oligomers interact with a mGluR5/cellular prion protein (PrPC) complex to disrupt physiological mGluR5 signal transduction. Aberrant mGluR5 signaling and associated synaptic failure are considered an emerging pathophysiological mechanism of Alzheimer's disease (AD). Therefore, mGluR5 represents an attractive therapeutic target for AD, and recent studies continue to validate the efficacy of various mGluR5 allosteric modulators in improving memory deficits and mitigating disease pathology. However, sex-specific differences in the pharmacology of mGluR5 and activation of noncanonical signaling downstream of the receptor suggest that its utility as a therapeutic target in female AD patients needs to be reconsidered.

VGLUT3 Deletion Rescues Motor Deficits and Neuronal Loss in the zQ175 Mouse Model of Huntington's Disease.

Huntington's disease (HD) is an autosomal-dominant neurodegenerative disease characterized by progressive motor and cognitive impairments, with no disease-modifying therapies yet available. HD pathophysiology involves evident impairment in glutamatergic neurotransmission leading to severe striatal neurodegeneration. The vesicular glutamate transporter-3 (VGLUT3) regulates the striatal network that is centrally affected by HD. Nevertheless, current evidence on the role of VGLUT3 in HD pathophysiology is lacking. Here, we crossed mice lacking Slc17a8 gene (VGLUT3 -/-) with heterozygous zQ175 knock-in mouse model of HD (zQ175:VGLUT3 -/-). Longitudinal assessment of motor and cognitive functions from 6 to 15 months of age reveals that VGLUT3 deletion rescues motor coordination and short-term memory deficits in both male and female zQ175 mice. VGLUT3 deletion also rescues neuronal loss likely via the activation of Akt and ERK1/2 in the striatum of zQ175 mice of both sexes. Interestingly, the rescue in neuronal survival in zQ175:VGLUT3 -/- mice is accompanied by a reduction in the number of nuclear mutant huntingtin (mHTT) aggregates with no change in the total aggregate levels or microgliosis. Collectively, these findings provide novel evidence that VGLUT3, despite its limited expression, can be a vital contributor to HD pathophysiology and a viable target for HD therapeutics.SIGNIFICANCE STATEMENT Dysregulation of the striatal network centrally contributes to the pathophysiology of Huntington's disease (HD). The atypical vesicular glutamate transporter-3 (VGLUT3) has been shown to regulate several major striatal pathologies, such as addiction, eating disorders, or L-DOPA-induced dyskinesia. Yet, our understanding of VGLUT3's role in HD remains unclear. We report here that deletion of the Slc17a8 (Vglut3) gene rescues the deficits in both motor and cognitive functions in HD mice of both sexes. We also find that VGLUT3 deletion activates neuronal survival signaling and reduces nuclear aggregation of abnormal huntingtin proteins and striatal neuron loss in HD mice. Our novel findings highlight the vital contribution of VGLUT3 in HD pathophysiology that can be exploited for HD therapeutic management.

Metabotropic Glutamate Receptor 2/3 Activation Improves Motor Performance and Reduces Pathology in Heterozygous zQ175 Huntington Disease Mice.

Huntington's disease (HD) is an autosomal dominant neurodegenerative disease that leads to progressive motor impairments with no available disease-modifying treatment. Current evidence indicates that exacerbated postsynaptic glutamate signaling in the striatum plays a key role in the pathophysiology of HD. However, it remains unclear whether reducing glutamate release can be an effective approach to slow the progression of HD. Here, we show that the activation of metabotropic glutamate receptors 2 and 3 (mGluR2/3), which inhibit presynaptic glutamate release, improves HD symptoms and pathology in heterozygous zQ175 knockin mice. Treatment of both male and female zQ175 mice with the potent and selective mGluR2/3 agonist LY379268 for either 4 or 8 weeks improves both limb coordination and locomotor function in all mice. LY379268 also reduces mutant huntingtin aggregate formation, neuronal cell death, and microglial activation in the striatum of both male and female zQ175 mice. The reduction in mutant huntingtin aggregates correlates with the activation of a glycogen synthase kinase 3ß-dependent autophagy pathway in male, but not female, zQ175 mice. Furthermore, LY379268 reduces both Akt and ERK1/2 phosphorylation in male zQ175 mice but increases both Akt and ERK1/2 phosphorylation in female zQ175 mice. Taken together, our results indicate that mGluR2/3 activation mitigates HD neuropathology in both male and female mice but is associated with the differential activation and inactivation of cell signaling pathways in heterozygous male and female zQ175 mice. This further highlights the need to take sex into consideration when developing future HD therapeutics. SIGNIFICANCE STATEMENT: The mGluR2/3 agonist LY379268 improves motor impairments and reduces pathology in male and female zQ175 Huntington's disease mice. The beneficial outcomes of LY379268 treatment in Huntington's disease mice were mediated by divergent cell signaling pathways in both sexes. We provide evidence that mGluR2/3 agonists can be repurposed for the treatment of Huntington's disease, and we emphasize the importance of investigating sex as a biological variable in preclinical disease-modifying studies.

The pleiotropic effects of antithrombotic drugs in the metabolic-cardiovascular-neurodegenerative disease continuum: impact beyond reduced clotting.

Antithrombotic drugs are widely used for primary and secondary prevention, as well as treatment of many cardiovascular disorders. Over the past few decades, major advances in the pharmacology of these agents have been made with the introduction of new drug classes as novel therapeutic options. Accumulating evidence indicates that the beneficial outcomes of some of these antithrombotic agents are not solely related to their ability to reduce thrombosis. Here, we review the evidence supporting established and potential pleiotropic effects of four novel classes of antithrombotic drugs, adenosine diphosphate (ADP) P2Y12-receptor antagonists, Glycoprotein IIb/IIIa receptor Inhibitors, and Direct Oral Anticoagulants (DOACs), which include Direct Factor Xa (FXa) and Direct Thrombin Inhibitors. Specifically, we discuss the molecular evidence supporting such pleiotropic effects in the context of cardiovascular disease (CVD) including endothelial dysfunction (ED), atherosclerosis, cardiac injury, stroke, and arrhythmia. Importantly, we highlight the role of DOACs in mitigating metabolic dysfunction-associated cardiovascular derangements. We also postulate that DOACs modulate perivascular adipose tissue inflammation and thus, may reverse cardiovascular dysfunction early in the course of the metabolic syndrome. In this regard, we argue that some antithrombotic agents can reverse the neurovascular damage in Alzheimer's and Parkinson's brain and following traumatic brain injury (TBI). Overall, we attempt to provide an up-to-date comprehensive review of the less-recognized, beneficial molecular aspects of antithrombotic therapy beyond reduced thrombus formation. We also make a solid argument for the need of further mechanistic analysis of the pleiotropic effects of antithrombotic drugs in the future.

Targeting Vesicular Glutamate Transporter Machinery: Implications on Metabotropic Glutamate Receptor 5 Signaling and Behavior.

Cross talk between both pre- and postsynaptic components of glutamatergic neurotransmission plays a crucial role in orchestrating a multitude of brain functions, including synaptic plasticity and motor planning. Metabotropic glutamate receptor (mGluR) 5 exhibits promising therapeutic potential for many neurodevelopmental and neurodegenerative disorders as a consequence of its modulatory control over diverse neuronal networks required for memory, motor coordination, neuronal survival, and differentiation. Given these crucial roles, mGluR5 signaling is under the tight control of glutamate release machinery mediated through vesicular glutamate transporters (VGLUTs) that ultimately dictate glutamatergic output. A particular VGLUT isoform, VGLUT3, exhibits an overlapping, but unique, distribution with mGluR5, and the dynamic cross talk between mGluR5 and VGLUT3 is key for the function of specific neuronal networks involved in motor coordination, emotions, and cognition. Thus, aberrant signaling of the VGLUT3-mGluR5 axis is linked to various pathologies including, but not limited to, Parkinson disease, anxiety disorders, and drug addiction. We argue that a comprehensive profiling of how coordinated VGLUT3-mGluR5 signaling influences overall glutamatergic neurotransmission is warranted. SIGNIFICANCE STATEMENT: Vesicular glutamate receptor (VGLUT) 3 machinery orchestrates glutamate release, and its distribution overlaps with metabotropic glutamate receptor (mGluR) 5 in regional brain circuitries, including striatum, hippocampus, and raphe nucleus. Therefore, VGLUT3-mGluR5 cross talk can significantly influence both physiologic and pathophysiologic glutamatergic neurotransmission. Pathological signaling of the VGLUT3-mGluR5 axis is linked to Parkinson disease, anxiety disorders, and drug addiction. However, it is also predicted to contribute to other motor and cognitive disorders.

Abnormal Rho-associated kinase activity contributes to the dysfunctional myogenic response of cerebral arteries in type 2 diabetes.

The structural and functional integrity of the brain, and therefore, cognition, are critically dependent on the appropriate control of blood flow within the cerebral circulation. Inadequate flow leads to ischemia, whereas excessive flow causes small vessel rupture and (or) blood-brain-barrier disruption. Cerebral blood flow is controlled through the interplay of several physiological mechanisms that regulate the contractile state of vascular smooth muscle cells (VSMCs) within the walls of cerebral resistance arteries and arterioles. The myogenic response of cerebral VSMCs is a key mechanism that is responsible for maintaining constant blood flow during variations in systemic pressure, i.e., flow autoregulation. Inappropriate myogenic control of cerebral blood flow is associated with, and prognostic of, neurological deterioration and poor outcome in patients with several conditions, including type 2 diabetes. Here, we review recent advances in our understanding of the role of inappropriate Rho-associated kinase activity as a cause of impaired myogenic regulation of cerebral arterial diameter in type 2 diabetes.

A M1 muscarinic acetylcholine receptor-specific positive allosteric modulator VU0486846 reduces neurogliosis in female Alzheimer's mice.

Alzheimer's disease (AD) is the most prevalent type of dementia, disproportionately affecting females, who make up nearly 60% of diagnosed cases. In AD patients, the accumulation of beta-amyloid (Aß) in the brain triggers a neuroinflammatory response driven by neuroglia, worsening the condition. We have previously demonstrated that VU0486846, an orally available positive allosteric modulator (PAM) targeting M1 muscarinic acetylcholine receptors, enhances cognitive function and reduces Aß pathology in female APPswe/PSEN1ΔE9 (APP/PS1) mice. However, it remained unclear whether these improvements were linked to a decrease in neuroglial activation. To investigate, we treated nine-month-old APP/PS1 and wildtype mice with VU0486846 for 8 weeks and analyzed brain slices for markers of microglial activation (ionized calcium binding adaptor molecule 1, Iba1) and astrocyte activation (Glial fibrillary acidic protein, GFAP). We find that VU0486846 reduces the presence of Iba1-positive microglia and GFAP-positive astrocytes in the hippocampus of female APP/PS1 mice and limits the recruitment of these cells to remaining Aß plaques. This study sheds light on an additional mechanism through which novel M1 mAChR PAMs exhibit disease-modifying effects by reducing neuroglial activation and underscore the potential of these ligands for the treatment of AD, especially in females.

The Role of Neuroglial Metabotropic Glutamate Receptors in Alzheimer's Disease.

Glutamate, the major excitatory neurotransmitter in the brain exerts its effects via both ionotropic glutamate receptors and metabotropic glutamate receptors (mGluRs). There are three subgroups of mGluRs, pre-synaptic Group II and Group III mGluRs and post-synaptic Group I mGluRs. mGluRs are ubiquitously expressed in the brain and their activation is poised upstream of a myriad of signaling pathways, resulting in their implication in the pathogenesis of various neurodegenerative diseases including, Alzheimer's Disease (AD). While the exact mechanism of AD etiology remains elusive, ß-amyloid (Aß) plaques and hyperphosphorylated tau tangles remain the histopathological hallmarks of AD. Though less electrically excitable, neuroglia are a major non-neuronal cell type in the brain and are composed of astrocytes, microglia, and oligodendrocytes. Astrocytes, microglia, and oligodendrocytes provide structural and metabolic support, active immune defence, and axonal support and sheathing, respectively. Interestingly, Aß and hyperphosphorylated tau are known to disrupt the neuroglial homeostasis in the brain, pushing them towards a more neurotoxic state. In this review, we discuss what is currently known regarding the expression patterns of various mGluRs in neuroglia and how Aß and tau alter the normal mGluR function in the neuroglia and contribute to the pathophysiology of AD.

Comparison of Huntington's disease phenotype progression in male and female heterozygous FDNQ175 mice.

Huntington's Disease (HD) is an inherited autosomal dominant neurodegenerative disorder that leads to progressive motor and cognitive impairment due to the expansion of a polyglutamine (CAG) repeat in the N-terminal region of the huntingtin (Htt) protein. The creation of HD mouse models represents a critical step in the research for HD treatment. Among the currently available HD mouse models, the zQ175 knock-in mouse line is the first to display robust disease phenotype on a heterozygous background. The newer FDNQ175 mouse model is derived from the zQ175 mouse line and presents a more aggressive phenotype. Moreover, increasing evidence has implicated sex as a contributing factor in the progression of HD symptoms. Here, we compared the progression of HD phenotypes in male and female heterozygous FDNQ175 mice. We found that both male and female heterozygous mice showed deficits in forelimb grip strength and cognition as early as 6 months of age. However, female FDNQ175 mice were less vulnerable to HD-associated decline in limb coordination and movement. Neither male nor female FDNQ175 mice exhibited reduced locomotor activity in the open field or exhibit consistent differences in anxiety at 6-12 months of age. Both male and female FDNQ175 mice exhibited increased numbers of huntingtin aggregates with age and 8-month-old female FDNQ175 mice had significantly more aggregates than their male counterparts. Taken together, our results provide further evidence that sex can influence the progression of HD phenotype in preclinical animal models and must be taken into consideration for future HD research.

Targeting mGluR group III for the treatment of neurodegenerative diseases.

Glutamate, an excitatory neurotransmitter, is essential for neuronal function, and it acts on ionotropic or metabotropic glutamate receptors (mGluRs). A disturbance in glutamatergic signaling is a hallmark of many neurodegenerative diseases. Developing disease-modifying treatments for neurodegenerative diseases targeting glutamate receptors is a promising avenue. The understudied group III mGluR 4, 6-8 are commonly found in the presynaptic membrane, and their activation inhibits glutamate release. Thus, targeted mGluRs therapies could aid in treating neurodegenerative diseases. This review describes group III mGluRs and their pharmacological ligands in the context of amyotrophic lateral sclerosis, Parkinson's, Alzheimer's, and Huntington's diseases. Attempts to evaluate the efficacy of these drugs in clinical trials are also discussed. Despite a growing list of group III mGluR-specific pharmacological ligands, research on the use of these drugs in neurodegenerative diseases is limited, except for Parkinson's disease. Future efforts should focus on delineating the contribution of group III mGluR to neurodegeneration and developing novel ligands with superior efficacy and a favorable side effect profile for the treatment of neurodegenerative diseases.

Targeting mGluR2/3 for treatment of neurodegenerative and neuropsychiatric diseases.

Glutamate is the primary excitatory neurotransmitter in the brain and plays critical roles in all aspects of neuronal function. Disruption of normal glutamate transmission has been implicated in a variety of neurodegenerative and neuropsychiatric diseases. Glutamate exerts its effect through ionotropic and metabotropic glutamate receptors (mGluRs). mGluR2 and mGluR3 are members of the Group II mGluR family and their activation leads to the inhibition of glutamate release from presynaptic nerve terminals and is also poised upstream of a myriad of signaling pathways in postsynaptic nerve terminals and neuroglia. Therefore, mGluR2 and mGluR3 have been considered as potential drug targets for the treatment of many neurological conditions and several compounds targeting these receptors have been developed. In this review, we discuss what is currently known regarding the contribution of mGluR2 and mGluR3 to the pathophysiology of some neurodegenerative and neuropsychiatric diseases including Amyotrophic lateral sclerosis, Alzheimer's disease, Huntington's disease, Parkinson's diseases, schizophrenia and depression as well as drug addiction. We then highlight the evidence supporting the use of various drugs including orthosteric and allosteric ligands acting on either mGluR2, mGluR3 or both for the management of these brain disorders.

VGLUT3 Ablation Differentially Modulates Glutamate Receptor Densities in Mouse Brain.

Type 3 vesicular glutamate transporter (VGLUT3) represents a unique modulator of glutamate release from both nonglutamatergic and glutamatergic varicosities within the brain. Despite its limited abundance, VGLUT3 is vital for the regulation of glutamate signaling and, therefore, modulates the activity of various brain microcircuits. However, little is known about how glutamate receptors are regulated by VGLUT3 across different brain regions. Here, we used VGLUT3 constitutive knock-out (VGLUT3-/-) mice and explored how VGLUT3 deletion influences total and cell surface expression of different ionotropic and metabotropic glutamate receptors. VGLUT3 deletion upregulated the overall expression of metabotropic glutamate receptors mGluR5 and mGluR2/3 in the cerebral cortex. In contrast, no change in the total expression of ionotropic NMDAR glutamate receptors were observed in the cerebral cortex of VGLUT3-/- mice. We noted significant reduction in cell surface levels of mGluR5, NMDAR2A, NMDAR2B, as well as reductions in dopaminergic D1 receptors and muscarinic M1 acetylcholine receptors in the hippocampus of VGLUT3-/- mice. Furthermore, mGluR2/3 total expression and mGluR5 cell surface levels were elevated in the striatum of VGLUT3-/- mice. Last, AMPAR subunit GluA1 was significantly upregulated throughout cortical, hippocampal, and striatal brain regions of VGLUT3-/- mice. Together, these findings complement and further support the evidence that VGLUT3 dynamically regulates glutamate receptor densities in several brain regions. These results suggest that VGLUT3 may play an intricate role in shaping glutamatergic signaling and plasticity in several brain areas.

Adipose tissue mitochondrial dysfunction and cardiometabolic diseases: On the search for novel molecular targets.

Cardiometabolic diseases present an escalating global health and economic burden. Such a surge is driven by epidemic prevalence rates of metabolic disorders, such as obesity and type 2 diabetes, and their associated cardiovascular complications, majorly contributing to morbidity and mortality. A fundamental challenge impeding the effective management and therapy of these complications is a lack of clear understanding of the molecular mechanisms underpinning disease initiation and progression. Over the past decade, a role for metabolic disease-associated adipose tissue dysfunction and inflammation in evoking cardiovascular and renal deterioration emerged, together with a growing recognition of the positive impact of pharmacological tools modulating adipose tissue function. Adipose tissue is a plastic endocrine organ whose homeostasis is essentially dependent on the intercellular communication of its comprising cellular components. Yet, despite being a principal regulator of adipose tissue metabolic activity, changes in aspects of adipose tissue mitochondrial biogenesis, dynamics, and bioenergetics in the context of cardiometabolic disorders have not garnered the necessary attention. Here, we gather the available evidence on the contribution of mitochondrial dysfunction to that of the adipose tissue in metabolic diseases, and to the ensuing cardiovascular deterioration. The involved molecular pathways are highlighted together with potential targets for intervention. The effects of several drug classes with known beneficial impact on adipose tissue remodeling and mitochondrial dysfunction in such a context are discussed. Finally, future research aspects in this domain are explored.

Metabotropic Glutamate Receptor 5 Antagonism Reduces Pathology and Differentially Improves Symptoms in Male and Female Heterozygous zQ175 Huntington's Mice.

Huntington's disease (HD) is an inherited autosomal dominant neurodegenerative disorder that leads to progressive motor and cognitive impairment. There are currently no available disease modifying treatments for HD patients. We have previously shown that pharmacological blockade of metabotropic glutamate receptor 5 (mGluR5) signaling rescues motor deficits, improves cognitive impairments and mitigates HD neuropathology in male zQ175 HD mice. Mounting evidence indicates that sex may influence HD progression and we have recently reported a sex-specific pathological mGluR5 signaling in Alzheimer's disease (AD) mice. Here, we compared the outcomes of treatment with the mGluR5 negative allosteric modulator CTEP (2-chloro-4-[2-[2,5-dimethyl-1-[4-(trifluoromethoxy)phenyl]imidazol-4-yl]ethynyl]pyridine) in both male and female symptomatic zQ175 mice. We found that female zQ175 mice required a longer treatment duration with CTEP than male mice to show improvement in their rotarod performance. Unlike males, chronic CTEP treatment did not improve the grip strength nor reverse the cognitive decline of female zQ175 mice. However, CTEP reduced the number of huntingtin aggregates, improved neuronal survival and decreased microglia activation in the striatum of both male and female zQ175 mice. Together, our results indicate that mGluR5 antagonism can reduce HD neuropathology in both male and female zQ175 HD mice, but sex has a clear impact on the efficacy of the treatment and must be taken into consideration for future HD drug development.

A positive allosteric modulator for the muscarinic receptor (M1 mAChR) improves pathology and cognitive deficits in female APPswe/PSEN1ΔE9 mice.

BACKGROUND AND PURPOSE: Alzheimer's disease (AD) is a neurodegenerative disease characterized by progressive cognitive decline, and women account for 60% of diagnosed cases. ß-Amyloid (Aß) oligomers are considered the principal neurotoxic species in AD brains. The M1 muscarinic ACh receptor (M1 mAChR) plays a key role in memory and learning. M1 mAChR agonists show pro-cognitive activity but cause many adverse off-target effects. A new orally bioavailable M1 mAChR positive allosteric modulator (PAM), VU0486846, is devoid of direct agonist activity or adverse effects but was not tested for disease-modifying efficacy in female AD mice. EXPERIMENTAL APPROACH: Nine-month-old female APPswe/PSEN1ΔE9 (APPswe) and wildtype mice were treated with VU0486846 in drinking water (10 mg·kg-1 ·day-1 ) for 4 or 8 weeks. Cognitive function of mice was assessed after treatment, and brains were harvested for biochemical and immunohistochemical assessment. KEY RESULTS: VU0486846 improved cognitive function of APPswe mice when tested in novel object recognition and Morris water maze. This was paralleled by a significant reduction in Aß oligomers and plaques and neuronal loss in the hippocampus. VU0486846 reduced Aß oligomer production in APPswe mice by increasing M1 mAChR expression and shifting the processing of amyloid precursor protein from amyloidogenic cleavage to non-amyloidogenic cleavage. Specifically, VU0486846 reduced the expression of ß-secretase 1 (BACE1), whereas it enhanced the expression of the α-secretase ADAM10 in APPswe hippocampus. CONCLUSION AND IMPLICATIONS: Using M1 mAChR PAMs can be a viable disease-modifying approach that should be exploited clinically to slow AD in women.

Early metabolic impairment as a contributor to neurodegenerative disease: Mechanisms and potential pharmacological intervention.

The metabolic syndrome comprises a family of clinical and laboratory findings, including insulin resistance, hyperglycemia, hypertriglyceridemia, low high-density lipoprotein cholesterol levels, and hypertension, in addition to central obesity. The syndrome confers a high risk of cardiovascular mortality. Indeed, metabolic dysfunction has been shown to cause a direct insult to smooth muscle and endothelial components of the vasculature, which leads to vascular dysfunction and hyperreactivity. This, in turn, causes cerebral vasoconstriction and hypoperfusion, eventually contributing to cognitive deficits. Moreover, the metabolic syndrome disrupts key homeostatic processes in the brain, including apoptosis, autophagy, and neurogenesis. Impairment of such processes in the context of metabolic dysfunction has been implicated in the pathogenesis of neurodegenerative diseases, including Alzheimer, Parkinson, and Huntington diseases. The aim of this review is to elucidate the role that the metabolic syndrome plays in the pathogenesis of the latter disorders, with a focus on the role of perivascular adipose inflammation in the peripheral-to-central transduction of the inflammatory insult. This review delineates common signaling pathways that contribute to these pathologies. Moreover, the role of therapeutic agents aimed at treating the metabolic syndrome, as well as their risk factors that interfere with the aforementioned pathways, are discussed as potential interventions for neurodegenerative diseases.

PPARγ dependence of cyclosporine-isoprenaline renovascular interaction: roles of nitric oxide synthase and heme oxygenase.

We previously showed that cyclosporine (CSA) impairs renal vasodilations caused by ß-adrenoceptor activation. This study investigated whether the peroxisome proliferator-activated receptor gamma (PPARγ) and related nitric oxide synthase (NOS)/heme oxygenase (HO) signaling mediates the CSA-ß-adrenoceptor interaction. The vasodilatory response elicited by a bolus injection of isoprenaline (1 µmole) in phenylephrine-preconstricted perfused kidneys of rats was reduced after prior infusion of zinc protoporphyrin IX (ZnPP, HO inhibitor) or GW9662 (PPARγ antagonist), suggesting the involvement of PPARγ and HO-derived CO in the isoprenaline response. In contrast, the inhibition of NOS activity by N-nitro-l-arginine methyl ester had no effect on isoprenaline responses. CSA (5 µM) significantly attenuated isoprenaline vasodilations, an effect that was abolished in the presence of GW9662 and accentuated by ZnPP. Also, supplementation with the PPARγ agonist pioglitazone or with l-arginine or hemin, substrates for NOS and HO, respectively, eliminated the unfavorable effect of CSA on isoprenaline vasodilations. The protection conferred by pioglitazone against CSA-evoked attenuation of isoprenaline vasodilations was maintained in N-nitro-l-arginine methyl ester-treated kidneys and disappeared after treatment with ZnPP or GW9662. In conclusion, the activation of the HO/CO/PPARγ cascade is probably the cellular mechanism that underlies the beneficial effect of pioglitazone on the CSA-isoprenaline interaction. Further, the facilitation of the HO/CO or NOS/NO pathway seems to offset this harmful effect of CSA.

Optineurin deletion disrupts metabotropic glutamate receptor 5-mediated regulation of ERK1/2, GSK3ß/ZBTB16, mTOR/ULK1 signaling in autophagy.

Optineurin (OPTN) is a multifunctional protein that mediates a network of cellular processes regulating membrane trafficking, inflammatory responses and autophagy. The OPTN-rich interactome includes Group I metabotropic glutamate receptors (mGluR1 and 5), members of the Gαq/11 protein receptor family. Recent evidence has shown that mGluR5, in addition to its canonical Gαq/11 protein-coupled signaling, regulates autophagic machinery via mTOR/ULK1 and GSK3ß/ZBTB16 pathways in both Alzheimer's and Huntington's disease mouse models. Despite its potential involvement, the role of OPTN in mediating mGluR5 downstream signaling cascades remains largely unknown. Here, we employed a CRISPR/Cas9 OPTN-deficient STHdhQ7/Q7 striatal cell line and global OPTN knockout mice to investigate whether Optn gene deletion alters both mGluR5 canonical and noncanonical signaling. We find that OPTN is required for mGluR5-activated Ca2+ flux and ERK1/2 signaling following receptor activation in STHdhQ7/Q7 cells and acute hippocampal slices. Deletion of OPTN impairs both GSK3ß/ZBTB16 and mTOR/ULK1 autophagic signaling in STHdhQ7/Q7 cells. Furthermore, mGluR5-dependent regulation of GSK3ß/ZBTB16 and mTOR/ULK1 autophagic signaling is impaired in hippocampal slices of OPTN knockout mice. Overall, we show that the crosstalk between OPTN and mGluR5 can have major implication on receptor signaling and therefore potentially contribute to the pathophysiology of neurodegenerative diseases.

Stromatoxin-sensitive, heteromultimeric Kv2.1/Kv9.3 channels contribute to myogenic control of cerebral arterial diameter.

Cerebral vascular smooth muscle contractility plays a crucial role in controlling arterial diameter and, thereby, blood flow regulation in the brain. A number of K(+) channels have been suggested to contribute to the regulation of diameter by controlling smooth muscle membrane potential (E(m)) and Ca(2+) influx. Previous studies indicate that stromatoxin (ScTx1)-sensitive, Kv2-containing channels contribute to the control of cerebral arterial diameter at 80 mmHg, but their precise role and molecular composition were not determined. Here, we tested if Kv2 subunits associate with 'silent' subunits from the Kv5, Kv6, Kv8 or Kv9 subfamilies to form heterotetrameric channels that contribute to control of diameter of rat middle cerebral arteries (RMCAs) over a range of intraluminal pressure from 10 to 100 mmHg. The predominant mRNAs expressed by RMCAs encode Kv2.1 and Kv9.3 subunits. Co-localization of Kv2.1 and Kv9.3 proteins at the plasma membrane of dissociated single RMCA myocytes was detected by proximity ligation assay. ScTx1-sensitive native current of RMCA myocytes and Kv2.1/Kv9.3 currents exhibited functional identity based on the similarity of their deactivation kinetics and voltage dependence of activation that were distinct from those of homomultimeric Kv2.1 channels. ScTx1 treatment enhanced the myogenic response of pressurized RMCAs between 40 and 100 mmHg, but this toxin also caused constriction between 10 and 40 mmHg that was not previously observed following inhibition of large conductance Ca(2+)-activated K(+) (BK(Ca)) and Kv1 channels. Taken together, this study defines the molecular basis of Kv2-containing channels and contributes to our understanding of the functional significance of their expression in cerebral vasculature. Specifically, our findings provide the first evidence of heteromultimeric Kv2.1/Kv9.3 channel expression in RMCA myocytes and their distinct contribution to control of cerebral arterial diameter over a wider range of E(m) and transmural pressure than Kv1 or BK(Ca) channels owing to their negative range of voltage-dependent activation.

Role of PPAR gamma/nitric oxide synthase signaling in the cyclosporine-induced attenuation of endothelium-dependent renovascular vasodilation.

Evidence from our laboratory and others suggests a negative effect for cyclosporine A (CSA) on renovascular reactivity. This study investigated the role of peroxisome proliferator-activated receptor gamma (PPAR gamma)/nitric oxide synthase (NOS) signaling in the CSA-induced attenuation of endothelium-dependent vasodilations in phenylephrine-preconstricted perfused kidneys of rats. Bolus injection of carbachol (4 micromoL) reduced the renal perfusion pressure with a peak depressor effect observed at 2 minutes. CSA (5 microM) infusion significantly attenuated the vasodilatory action of carbachol. The specificity of this interaction was verified by the lack of effect of CSA on renal vasodilation caused by papaverine (50 nmol). The carbachol-induced renal vasodilations were also reduced after infusion of N-nitro-L-arginine methyl ester (L-NAME, NOS inhibitor, 100 microM) or 2-chloro-5-nitro-N-phenylbenzamide (GW9662, PPAR gamma antagonist, 1 microM). The attenuation of carbachol vasodilation by CSA was abolished in presence of L-arginine or L-NAME in contrast to no effect for GW9662. Pioglitazone (PPAR gamma agonist, 10 microM) abolished the CSA-induced attenuation of carbachol responses, an effect that was not manifest in presence of GW9662 or l-NAME. These findings suggest that PPAR gamma act tonically to facilitate renovascular dilatory response to endothelial muscarinic receptor activation. More importantly, NOS signaling downstream of PPAR gamma mediates, at least partly, the inhibitory effect of CSA on carbachol vasodilations.