Lipocalin-type prostaglandin D synthase regulates light-induced phase advance of the central circadian rhythm in mice.

We previously showed that mice lacking pituitary adenylate cyclase-activating polypeptide (PACAP) exhibit attenuated light-induced phase shift. To explore the underlying mechanisms, we performed gene expression analysis of laser capture microdissected suprachiasmatic nuclei (SCNs) and found that lipocalin-type prostaglandin (PG) D synthase (L-PGDS) is involved in the impaired response to light stimulation in the late subjective night in PACAP-deficient mice. L-PGDS-deficient mice also showed impaired light-induced phase advance, but normal phase delay and nonvisual light responses. Then, we examined the receptors involved in the response and observed that mice deficient for type 2 PGD2 receptor DP2/CRTH2 (chemoattractant receptor homologous molecule expressed on Th2 cells) show impaired light-induced phase advance. Concordant results were observed using the selective DP2/CRTH2 antagonist CAY10471. These results indicate that L-PGDS is involved in a mechanism of light-induced phase advance via DP2/CRTH2 signaling.

Unbiased compound screening with a reporter gene assay highlights the role of p13 in the cardiac cellular stress response.

We recently showed that a 13-kDa protein (p13), the homolog protein of formation of mitochondrial complex V assembly factor 1 in yeast, acts as a potential protective factor in pancreatic islets under diabetes. Here, we aimed to identify known compounds regulating p13 mRNA expression to obtain therapeutic insight into the cellular stress response. A luciferase reporter system was developed using the putative promoter region of the human p13 gene. Overexpression of peroxisome proliferator-activated receptor gamma coactivator 1α, a master player regulating mitochondrial metabolism, increased both reporter activity and p13 expression. Following unbiased screening with 2320 known compounds in HeLa cells, 12 pharmacological agents (including 8 cardiotonics and 2 anthracyclines) that elicited >2-fold changes in p13 mRNA expression were identified. Among them, four cardiac glycosides decreased p13 expression and concomitantly elevated cellular oxidative stress. Additional database analyses showed highest p13 expression in heart, with typically decreased expression in cardiac disease. Accordingly, our results illustrate the usefulness of unbiased compound screening as a method for identifying novel functional roles of unfamiliar genes. Our findings also highlight the importance of p13 in the cellular stress response in heart.

High-Speed and Scalable Whole-Brain Imaging in Rodents and Primates.

Subcellular resolution imaging of the whole brain and subsequent image analysis are prerequisites for understanding anatomical and functional brain networks. Here, we have developed a very high-speed serial-sectioning imaging system named FAST (block-face serial microscopy tomography), which acquires high-resolution images of a whole mouse brain in a speed range comparable to that of light-sheet fluorescence microscopy. FAST enables complete visualization of the brain at a resolution sufficient to resolve all cells and their subcellular structures. FAST renders unbiased quantitative group comparisons of normal and disease model brain cells for the whole brain at a high spatial resolution. Furthermore, FAST is highly scalable to non-human primate brains and human postmortem brain tissues, and can visualize neuronal projections in a whole adult marmoset brain. Thus, FAST provides new opportunities for global approaches that will allow for a better understanding of brain systems in multiple animal models and in human diseases.

Prostaglandin D2 signaling mediated by the CRTH2 receptor is involved in MK-801-induced cognitive dysfunction.

Chemoattractant receptor-homologous molecule expressed on T helper type 2 cells (CRTH2), which is a second receptor for prostaglandin (PG) D2, is involved in inflammatory responses in peripheral tissue; however, its role in cognitive function remains unclear. Here, we demonstrate that CRTH2 is involved in cognitive function using a well-established animal model of cognitive dysfunction induced by MK-801, an N-methyl-d-aspartate receptor antagonist. Genetic deletion and pharmacological inhibition of CRTH2 suppressed MK-801-induced cognitive dysfunction. Pharmacological inhibition of cyclooxygenase-1, a rate-limiting enzyme in PG synthesis, also suppressed MK-801-induced cognitive dysfunction. Moreover, an MK-801-induced increase in c-Fos expression in the paraventricular nucleus (PVN) was abolished in the CRTH2-deficient mice. Together, these results suggest that PGD2-CRTH2 signaling is involved in both MK-801-induced cognitive dysfunction and neuronal activity regulation in the PVN. Furthermore, genetic association studies suggest that CRTH2 is weakly associated with cognitive function in humans. Our study provides evidence that PGD2-CRTH2 signaling is involved in cognitive function and may represent a potential therapeutic target for cognitive dysfunction in patients with psychiatric disorders.

PACAP suppresses dry eye signs by stimulating tear secretion.

Dry eye syndrome is caused by a reduction in the volume or quality of tears. Here, we show that pituitary adenylate cyclase-activating polypeptide (PACAP)-null mice develop dry eye-like symptoms such as corneal keratinization and tear reduction. PACAP immunoreactivity is co-localized with a neuronal marker, and PACAP receptor (PAC1-R) immunoreactivity is observed in mouse infraorbital lacrimal gland acinar cells. PACAP eye drops stimulate tear secretion and increase cAMP and phosphorylated (p)-protein kinase A levels in the infraorbital lacrimal glands that could be inhibited by pre-treatment with a PAC1-R antagonist or an adenylate cyclase inhibitor. Moreover, these eye drops suppress corneal keratinization in PACAP-null mice. PACAP eye drops increase aquaporin 5 (AQP5) levels in the membrane and pAQP5 levels in the infraorbital lacrimal glands. AQP5 siRNA treatment of the infraorbital lacrimal gland attenuates PACAP-induced tear secretion. Based on these results, PACAP might be clinically useful to treat dry eye disorder.

Endothelial Angiogenesis and Barrier Function in Response to Thrombin Require Ca2+ Influx through the Na+/Ca2+ Exchanger.

Thrombin acts on the endothelium by activating protease-activated receptors (PARs). The endothelial thrombin-PAR system becomes deregulated during pathological conditions resulting in loss of barrier function and a pro-inflammatory and pro-angiogenic endothelial phenotype. We reported recently that the ion transporter Na(+)/Ca(2+) exchanger (NCX) operating in the Ca(2+)-influx (reverse) mode promoted ERK1/2 activation and angiogenesis in vascular endothelial growth factor-stimulated primary human vascular endothelial cells. Here, we investigated whether Ca(2+) influx through NCX was involved in ERK1/2 activation, angiogenesis, and endothelial barrier dysfunction in response to thrombin. Reverse-mode NCX inhibitors and RNAi-mediated NCX1 knockdown attenuated ERK1/2 phosphorylation in response to thrombin or an agonist of PAR-1, the main endothelial thrombin receptor. Conversely, promoting reverse-mode NCX by suppressing Na(+)-K(+)-ATPase activity enhanced ERK1/2 activation. Reverse-mode NCX inhibitors and NCX1 siRNA suppressed thrombin-induced primary human vascular endothelial cell angiogenesis, quantified as proliferation and tubular differentiation. Reverse-mode NCX inhibitors or NCX1 knockdown preserved barrier integrity upon thrombin stimulation in vitro. Moreover, the reverse-mode NCX inhibitor SEA0400 suppressed Evans' blue albumin extravasation to the lung and kidneys and attenuated edema formation and ERK1/2 activation in the lungs of mice challenged with a peptide activator of PAR-1. Mechanistically, thrombin-induced ERK1/2 activation required NADPH oxidase 2-mediated reactive oxygen species (ROS) production, and reverse-mode NCX inhibitors and NCX1 siRNA suppressed thrombin-induced ROS production. We propose that reverse-mode NCX is a novel mechanism contributing to thrombin-induced angiogenesis and hyperpermeability by mediating ERK1/2 activation in a ROS-dependent manner. Targeting reverse-mode NCX could be beneficial in pathological conditions involving unregulated thrombin signaling.

p13 overexpression in pancreatic ß-cells ameliorates type 2 diabetes in high-fat-fed mice.

We examined the pancreatic function of p13 encoded by 1110001J03Rik, whose expression is decreased in pancreatic islets in high-fat-fed diabetic mice, by generating transgenic mice overexpressing p13 (p13-Tg) in pancreatic ß-cells. p13-Tg mice showed normal basal glucose metabolism; however, under high-fat feeding, these animals showed augmented glucose-induced first-phase and total insulin secretion, improved glucose disposal, greater islet area and increased mitotic insulin-positive cells. In addition, high-fat diet-induced 4-hydroxynonenal immunoreactivity, a reliable marker and causative agent of lipid peroxidative stress, was significantly decreased in p13-Tg mouse islets. These results indicate that p13 is a novel pancreatic factor exerting multiple beneficial effects against type 2 diabetes.

PACAP enhances axon outgrowth in cultured hippocampal neurons to a comparable extent as BDNF.

Pituitary adenylate cyclase-activating polypeptide (PACAP) exerts neurotrophic activities including modulation of synaptic plasticity and memory, hippocampal neurogenesis, and neuroprotection, most of which are shared with brain-derived neurotrophic factor (BDNF). Therefore, the aim of this study was to compare morphological effects of PACAP and BDNF on primary cultured hippocampal neurons. At days in vitro (DIV) 3, PACAP increased neurite length and number to similar levels by BDNF, but vasoactive intestinal polypeptide showed much lower effects. In addition, PACAP increased axon, but not dendrite, length, and soma size at DIV 3 similarly to BDNF. The PACAP antagonist PACAP6-38 completely blocked the PACAP-induced increase in axon, but not dendrite, length. Interestingly, the BDNF-induced increase in axon length was also inhibited by PACAP6-38, suggesting a mechanism involving PACAP signaling. K252a, a TrkB receptor inhibitor, inhibited axon outgrowth induced by PACAP and BDNF without affecting dendrite length. These results indicate that in primary cultured hippocampal neurons, PACAP shows morphological actions via its cognate receptor PAC1, stimulating neurite length and number, and soma size to a comparable extent as BDNF, and that the increase in total neurite length is ascribed to axon outgrowth.

CRTH2, a prostaglandin D2 receptor, mediates depression-related behavior in mice.

Depression is a complex neuropsychiatric disorder with an unclear molecular etiology. Inflammatory cytokines and molecular intermediates (including prostaglandins) are suggested to be involved in depression; however, the roles of prostaglandins and their respective receptors are largely unknown in depression. Using genetic and pharmacological approaches, we show here that chemoattractant receptor-homologous molecule expressed on T helper type 2 cells (CRTH2), a second receptor for prostaglandin D2 (PGD2), mediates depression-related behavior in mice. CRTH2-deficient (CRTH2(-/-)) mice showed antidepressant-like activity in a chronic corticosterone treatment-induced depression. Consistent with this observation, the pharmacological inhibition of CRTH2 via the clinically available drug ramatroban also rescued abnormal social interaction and depression-related behavior in well-established models, including chronic corticosterone-, lipopolysaccharide-, and tumor-induced pathologically relevant depression models. Importantly, chronic stress via corticosterone treatment increased mRNA levels in PGD2-producing enzymes, such as cyclooxygenase-2 and lipocalin-type PGD2 synthase, in the brain. Furthermore, the activity of the hippocampal noradrenergic system but not the dopaminergic or serotonergic systems was increased in CRTH2(-/-) mice. Together with the observation that untreated CRTH2(-/-) mice showed antidepressant-like activity in the forced swim test, these results provide evidence that central CRTH2-mediated signaling is critically involved in depression-related behavior.

Simultaneous neuron- and astrocyte-specific fluorescent marking.

Systematic and simultaneous analysis of multiple cell types in the brain is becoming important, but such tools have not yet been adequately developed. Here, we aimed to generate a method for the specific fluorescent labeling of neurons and astrocytes, two major cell types in the brain, and we have developed lentiviral vectors to express the red fluorescent protein tdTomato in neurons and the enhanced green fluorescent protein (EGFP) in astrocytes. Importantly, both fluorescent proteins are fused to histone 2B protein (H2B) to confer nuclear localization to distinguish between single cells. We also constructed several expression constructs, including a tandem alignment of the neuron- and astrocyte-expression cassettes for simultaneous labeling. Introducing these vectors and constructs in vitro and in vivo resulted in cell type-specific and nuclear-localized fluorescence signals enabling easy detection and distinguishability of neurons and astrocytes. This tool is expected to be utilized for the simultaneous analysis of changes in neurons and astrocytes in healthy and diseased brains.

Comparative gene expression profiles in pancreatic islets associated with agouti yellow mutation and PACAP overexpression in mice.

In diabetes mellitus, pituitary adenylate cyclase-activating polypeptide (PACAP) has insulinotropic and glucose-lowering properties. We previously demonstrated that transgenic mice overexpressing PACAP in pancreatic ß-cells (PACAP-Tg) show attenuated pancreatic islet hyperplasia and hyperinsulinemia in type 2 diabetic models. To explore the underlying mechanisms, here we crossed PACAP-Tg mice with lethal yellow agouti (KKAy) diabetic mice, and performed gene chip analysis of laser capture microdissected pancreatic islets from four F1 offspring genotypes (wild-type, PACAP-Tg, KKAy, and PACAP-Tg:KKAy). We identified 1371 probes with >16-fold differences between at least one pair of genotypes, and classified the probes into five clusters with characteristic expression patterns. Gene ontology enrichment analysis showed that genes involved in the terms ribosome and intracellular organelles such as ribonucleoprotein complex, mitochondrion, and chromosome organization were significantly enriched in clusters characterized by up-regulated genes in PACAP-Tg:KKAy mice compared with KKAy mice. These results may provide insight into the mechanisms of diabetes that accompany islet hyperplasia and amelioration by PACAP.

Identification of the role of bone morphogenetic protein (BMP) and transforming growth factor-ß (TGF-ß) signaling in the trajectory of serotonergic differentiation in a rapid assay in mouse embryonic stem cells in vitro.

The mechanism by which extracellular molecules control serotonergic cell fate remains elusive. Recently, we showed that noggin, which inactivates bone morphogenetic proteins (BMPs), induces serotonergic differentiation of mouse embryonic (ES) and induced pluripotent stem cells with coordinated gene expression along the serotonergic lineage. Here, we created a rapid assay for serotonergic induction by generating knock-in ES cells expressing a naturally secreted Gaussia luciferase driven by the enhancer of Pet-1/Fev, a landmark of serotonergic differentiation. Using these cells, we performed candidate-based screening and identified BMP type I receptor kinase inhibitors LDN-193189 and DMH1 as activators of luciferase. LDN-193189 induced ES cells to express the genes encoding Pet-1, tryptophan hydroxylase 2, and the serotonin transporter, and increased serotonin release without altering dopamine release. In contrast, TGF-ß receptor inhibitor SB-431542 selectively inhibited serotonergic differentiation, without changing overall neuronal differentiation. LDN-193189 inhibited expression of the BMP signaling target gene Id, and induced the TGF-ß target gene Lefty, whereas the opposite effect was observed with SB-431542. This study thus provides a new tool to investigate serotonergic differentiation and suggests that inhibition of BMP type I receptors and concomitant activation of TGF-ß receptor signaling are implicated in serotonergic differentiation. Candidate-based screening for serotonergic induction using a rapid assay in mouse embryonic stem cells revealed that the bone morphogenetic protein (BMP) type I receptor kinase inhibitors selectively induce serotonergic differentiation, whereas the TGF-ß receptor inhibitor SB-431542 inhibits the differentiation. These results suggest that inhibition of BMP type I receptors and concomitant activation of transforming growth factor-ß (TGF-ß) receptor signaling are involved in the early trajectory of serotonergic differentiation.

An enriched environment ameliorates memory impairments in PACAP-deficient mice.

We previously found that juvenile pituitary adenylate cyclase-activating polypeptide (PACAP)-knockout (PACAP(-/-)) mice reared in an enriched environment (EE) for 4 weeks showed attenuated hyperactivity, jumping behavior, impairments in social interaction, and depression-like behavior. The present study examined the effects of EE on memory function and memory-related protein levels in PACAP(-/-) mice. Eight-week-old PACAP(-/-) mice displayed fear memory dysfunction in a contextual fear conditioning test and cognitive impairments in a novel object recognition test. Rearing of 4-week-old PACAP(-/-) mice in an EE for 4 weeks ameliorated these memory impairments. The beneficial effects of EE were also observed 2 weeks after a return to housing in a standard environment (SE). This suggests that the effects of EE on impaired memory are long-lasting. In both wild-type and PACAP(-/-) mice, EE increased the protein levels of the NMDA receptor NR2B subunit, phospho-ERK, phospho-CaMKII, and brain-derived neurotrophic factor (BDNF) in the hippocampus, and decreased neurotrophin-3 levels, whereas it did not affect nerve growth factor and glial cell-derived neurotrophic factor levels. Increased levels of NR2B, phospho-ERK, phospho-CaMKII and BDNF were not observed 2 weeks after a return to housing in a SE. These findings suggest that living in an EE engenders long-lasting reductions in memory impairments in PACAP(-/-) mice. The present study also implies that increases in hippocampal memory-related protein and BDNF levels are responsible for the beneficial effects of an EE, but not for the maintenance of these effects.

The effect of high [K(+)]o on spontaneous Ca(2+) waves in freshly isolated interstitial cells of Cajal from the rabbit urethra.

Interstitial cells of Cajal (ICC) act as putative pacemaker cells in the rabbit urethra. Pacemaker activity in ICC results from spontaneous global Ca(2+) waves that can be increased in frequency by raising external [K(+)]. The purpose of this study was to elucidate the mechanism of this response. Intracellular [Ca(2+)] was measured in fluo-4-loaded smooth muscle cells (SMCs) and ICC using a Nipkow spinning disk confocal microscope. Increasing [K(+)]o to 60 mmol/L caused an increase in [Ca(2+)]i accompanied by contraction in SMCs. Raising [K(+)]o did not cause contraction in ICC, but the frequency of firing of spontaneous calcium waves increased. Reducing [Ca(2+)]o to 0 mmol/L abolished the response in both cell types. Nifedipine of 1 µmol/L blocked the response of SMC to high [K(+)]o, but did not affect the increase in firing in ICC. This latter effect was blocked by 30 µmol/L NiCl2 but not by the T-type Ca(2+) channel blocker mibefradil (300 nmol/L). However, inhibition of Ca(2+) influx via reverse-mode sodium/calcium exchange (NCX) using either 1 µmol/L SEA0400 or 5 µmol/L KB-R7943 did block the effect of high [K(+)]o on ICC. These data suggest that high K(+) solution increases the frequency of calcium waves in ICC by increasing Ca(2+) influx through reverse-mode NCX.

Increased behavioral and neuronal responses to a hallucinogenic drug in PACAP heterozygous mutant mice.

Accumulating evidence from human genetic studies implicates the pituitary adenylate cyclase-activating polypeptide (PACAP) gene as a risk factor for psychiatric disorders, including schizophrenia and stress-related diseases. Mice with homozygous disruption of the PACAP gene display profound behavioral and neurological abnormalities that are ameliorated with the atypical antipsychotic and dopamine D2 and serotonin (5-HT)2 antagonist risperidone and the 5-HT2 receptor antagonist ritanserin; however, the underlying mechanisms remain unknown. Here, we investigated if PACAP heterozygous mutant (PACAP(+/-)) mice, which appear behaviorally normal, are vulnerable to aversive stimuli. PACAP(+/-) mice were administered a 5-HT2 receptor agonist, (±)-2,5-dimethoxy-4-iodoamphetamine (DOI), a hallucinogenic drug, and their responses were compared with the littermate wild-type mice. After DOI injection, PACAP(+/-) mice showed increased head-twitch responses, while their behavior was normal after saline. DOI induced deficits in sensorimotor gating, as determined by prepulse inhibition, specifically in PACAP(+/-) mice. However, other 5-HT2 receptor-dependent responses, such as corticosterone release and hypothermia, were similarly observed in PACAP(+/-) and wild-type mice. c-Fos expression analysis, performed in various brain regions, revealed that the DOI-induced increase in the number of c-Fos-positive cells was more pronounced in 5-HT2A receptor-negative cells in the somatosensory cortex in PACAP(+/-) mice compared with wild-type mice. These results indicate that PACAP(+/-) mice exhibit specific vulnerability to DOI-induced deficits in cortical sensory function, such as exaggerated head-twitch responses and sensorimotor gating deficits. Our findings provide insight into the neural mechanisms underlying impaired behavioral responses in which 5-HT2 receptors are implicated.

Central CRTH2, a second prostaglandin D2 receptor, mediates emotional impairment in the lipopolysaccharide and tumor-induced sickness behavior model.

Chemoattractant receptor-homologous molecule expressed on T helper type 2 cells (CRTH2) is a second prostaglandin D2 receptor involved in mediating the allergic response; however, its central function is not yet known. Here, we demonstrate that central CRTH2 mediates emotional impairment. Lipopolysaccharide (LPS)-induced decreases in social interaction and novel exploratory behavior were observed in wild-type (CRTH2(+/+)) mice but not CRTH2-deficient (CRTH2(-/-)) mice, but both genotypes showed hypolocomotion and anorexia following LPS injection. Tumor (colon 26) inoculation, a more pathologically relevant model, induced decreases in social interaction and novel exploratory behavior in CRTH2(+/+), but not CRTH2(-/-) mice. In addition, the CRTH2 antagonists including clinically available ramatroban reversed impaired social interaction and novel exploratory behavior after either LPS or tumor inoculation in CRTH2(+/+) mice. Finally, LPS-induced c-Fos expression in the hypothalamic paraventricular nucleus (PVN) and central amygdala (CeA) was selectively abolished in CRTH2(-/-) mice. These results show that CRTH2 participates in LPS-induced emotional changes and activation in the PVN and CeA. Our study provides the first evidence that central CRTH2 regulates specific emotional behaviors, and that CRTH2 antagonism has potential as a therapeutic target for behavioral symptoms associated with tumors and infectious diseases.

Stimulation of Na(+)/H(+) exchanger isoform 1 promotes microglial migration.

Regulation of microglial migration is not well understood. In this study, we proposed that Na(+)/H(+) exchanger isoform 1 (NHE-1) is important in microglial migration. NHE-1 protein was co-localized with cytoskeletal protein ezrin in lamellipodia of microglia and maintained its more alkaline intracellular pH (pHi). Chemoattractant bradykinin (BK) stimulated microglial migration by increasing lamellipodial area and protrusion rate, but reducing lamellipodial persistence time. Interestingly, blocking NHE-1 activity with its potent inhibitor HOE 642 not only acidified microglia, abolished the BK-triggered dynamic changes of lamellipodia, but also reduced microglial motility and microchemotaxis in response to BK. In addition, NHE-1 activation resulted in intracellular Na(+) loading as well as intracellular Ca(2+) elevation mediated by stimulating reverse mode operation of Na(+)/Ca(2+) exchange (NCXrev). Taken together, our study shows that NHE-1 protein is abundantly expressed in microglial lamellipodia and maintains alkaline pHi in response to BK stimulation. In addition, NHE-1 and NCXrev play a concerted role in BK-induced microglial migration via Na(+) and Ca(2+) signaling.

PACAP Inhibits ß-cell Mass Expansion in a Mouse Model of Type II Diabetes: Persistent Suppressive Effects on Islet Density.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a potent insulinotropic G-protein-coupled receptor ligand, for which morphoregulative roles in pancreatic islets have recently been suggested. Here, we evaluated the effects of pancreatic overexpression of PACAP on morphometric changes of islets in a severe type II diabetes model. Following cross-breeding of obese-diabetic model KKA(y) mice with mice overexpressing PACAP in their pancreatic ß-cells, the resulting KKA(y) mice with or without PACAP transgene (PACAP/+:A(y)/+ or A(y)/+ mice) were fed with a high-fat diet up to the age of 11 months. Pancreatic sections from 5- to 11-month-old littermates were examined. Histomorphometric analyses revealed significant suppression of islet mass expansion in PACAP/+:A(y)/+ mice compared with A(y)/+ mice at 11 months, but no significant difference between PACAP/+ and +/+ (wild-type) mice, as previously reported. The suppressed islet mass in PACAP/+:A(y)/+ mice was due to a decrease in islet density but not islet size. In addition, the density of tiny islets (<0.001 mm(2)) and of insulin-positive clusters in ductal structures were markedly decreased in PACAP/+:A(y)/+ mice compared with A(y)/+ mice at 5 months of age. In contrast, PACAP overexpression caused no significant effects on the level of aldehyde-fuchsin reagent staining (a measure of ß-cell granulation) or the volume and localization of glucagon-positive cells in the pancreas. These results support previously reported inhibitory effects of PACAP on pancreatic islet mass expansion, and suggest it has persistent suppressive effects on pancreatic islet density which may be related with ductal cell-associated islet neogenesis in type II diabetes.

Effects of SEA0400 on arrhythmogenicity in a Langendorff-perfused 1-month myocardial infarction rabbit model.

BACKGROUND: The effects of SEA0400, a Na(+) /Ca(2+) exchanger (NCX) blocker, on dynamic factors and arrhythmogenic alternans in 1-month myocardial infarction (MI) hearts remain unknown. METHODS: Simultaneous voltage and intracellular Ca(2+) (Cai ) optical mapping was performed in 12 rabbit hearts with MI for 1 month and six normal rabbit hearts as control. Western-blot studies were performed in both groups in an additional six hearts for each. Action potential duration (APD) restitution was constructed and arrhythmogenic alternans was induced by dynamic pacing. SEA0400 (0.03, 3 µM) was administered after baseline studies. RESULTS: SEA0400 suppressed pacing-induced ventricular premature beats in a concentration-dependent manner. SEA0400 at 0.03 µM steepened APD restitution slopes and enhanced spatially discordant alternans (SDA), which became insignificant at 3 µM. The VF inducibility was seven of nine at baseline, nine of nine at 0.03 µM SEA0400, and five of nine at 3 µM SEA0400 (P = NS). Significant upregulation of NCX in the remote but not periinfarct zone and less degree downregulation of DHP1α in the remote versus periinfarct zone may play a role in enhancing SDA induction by SEA0400 in 1-month MI hearts. CONCLUSIONS: In 1-month MI hearts, SEA0400 suppresses pacing-induced ventricular premature beats, but also is proarrhythmic by steepening APD restitution and enhancing SDA via NCX inhibition. Heterogeneous upregulation of NCX and downregulation of DHP1α may contribute to SDA augmentation by SEA0400 in this model. The insignificant effect of SEA0400 on VF inducibility suggests that suppression of both reentry and triggered activity is required to suppress VF induction in this model.

The selective metabotropic glutamate 2/3 receptor agonist MGS0028 reverses psychomotor abnormalities and recognition memory deficits in mice lacking the pituitary adenylate cyclase-activating polypeptide.

Previous studies suggest that metabotropic glutamate 2/3 receptors are involved in psychiatric disorders. In this study, we examined the effects of the selective metabotropic glutamate 2/3 (mGlu2/3) receptor agonist MGS0028 on behavioral abnormalities in mice lacking the pituitary adenylate cyclase-activating polypeptide (PACAP), an experimental model of psychiatric disorders such as schizophrenia and attention-deficit/hyperactivity disorder. We found that PACAP-deficient mice showed impairments in the novel object recognition test and these impairments were improved by MGS0028 (0.1 mg/kg). Similarly, MGS0028 improved hyperactivity and jumping behaviors, but did not reverse increased immobility times in the forced swim test in PACAP-deficient mice. These results suggest that MGS0028 may be a potential, novel treatment for psychiatric disorders.