Subunit Of Voltage-gated Calcium Channels Research Articles

A New Zebrafish Model for CACNA2D4-Dysfunction.

Mutations in CACNA2D4, encoding the α2δ4 subunit of retinal voltage-gated calcium channels (Cav), cause a rare type of retinal dysfunction in human, mainly affecting cone vision. Here, we investigate the role of CACNA2D4 in targeting of Cav, its influence on cone-mediated signal transmission, and the cellular and subcellular changes upon loss of α2δ4 by exploiting the advantages of the cone-dominant zebrafish as model system. We identified two zebrafish CACNA2D4 paralogs (cacna2d4a and cacna2d4b), analyzed their expression by RNA in situ hybridization and introduced truncating frameshift mutations through CRISPR/Cas9-mediated mutagenesis. We analyzed retinal function and morphology of the single and double mutant lines by electroretinography, immunohistochemistry, light- and electron microscopy. Knockout of cacna2d4b reduces the expression of Cacna1fa, the pore-forming subunit of retinal Cav1.4, whereas loss of cacna2d4a did not. Only knockout of both paralogs impaired cone-mediated ERG b-wave amplitude. The number of "floating" ribbons is increased in double-KO, while retinal morphology and expression of postsynaptic mGluR6b remain largely unaffected. Both Cacna1fa and Ribeyeb show ectopic punctate expression in cacna2d4b-KO and double-KO photoreceptors. We find that increasing the expression of Cav at the synaptic membrane is an evolutionarily conserved function of Cacna2d4b. Yet, since both paralogs participate in cone synaptic transmission, we propose partial subfunctionalization in zebrafish. Similar to human patients, our double KO zebrafish model shows mild cone dysfunction, which was not associated with signs of retinal degeneration. Therefore, cacna2d4-KO zebrafish is a suitable model to study the pathophysiological mechanisms underlying CACNA2D4 dysfunction in human.

Gabapentinoid treatment promotes corticospinal plasticity and regeneration following murine spinal cord injury.

Axon regeneration failure causes neurological deficits and long-term disability after spinal cord injury (SCI). Here, we found that the α2δ2 subunit of voltage-gated calcium channels negatively regulates axon growth and regeneration of corticospinal neurons, the cells that originate the corticospinal tract. Increased α2δ2 expression in corticospinal neurons contributed to loss of corticospinal regrowth ability during postnatal development and after SCI. In contrast, α2δ2 pharmacological blockade through gabapentin administration promoted corticospinal structural plasticity and regeneration in adulthood. Using an optogenetic strategy combined with in vivo electrophysiological recording, we demonstrated that regenerating corticospinal axons functionally integrate into spinal circuits. Mice administered gabapentin recovered upper extremity function after cervical SCI. Importantly, such recovery relies on reorganization of the corticospinal pathway, as chemogenetic silencing of injured corticospinal neurons transiently abrogated recovery. Thus, targeting α2δ2 with a clinically relevant treatment strategy aids repair of motor circuits after SCI.

RBM20 Regulates CaV1.2 Surface Expression by Promoting Exon 9* Inclusion of CACNA1C in Neonatal Rat Cardiomyocytes.

The CACNA1C gene encodes for the CaV1.2 protein, which is the pore subunit of cardiac l-type voltage-gated calcium (Ca2+) channels (l-channels). Through alternative splicing, CACNA1C encodes for various CaV1.2 isoforms with different electrophysiological properties. Splice variants of CaV1.2 are differentially expressed during heart development or pathologies. The molecular mechanisms of CACNA1C alternative splicing still remain incompletely understood. RNA sequencing analysis has suggested that CACNA1C is a potential target of the splicing factor RNA-binding protein motif 20 (RBM20). Here, we aimed at elucidating the role of RBM20 in the regulation of CACNA1C alternative splicing. We found that in neonatal rat cardiomyocytes (NRCMs), RBM20 overexpression promoted the inclusion of CACNA1C’s exon 9*, whereas the skipping of exon 9* occurred upon RBM20 siRNA knockdown. The splicing of other known alternative exons was not altered by RBM20. RNA immunoprecipitation suggested that RBM20 binds to introns flanking exon 9*. Functionally, in NRCMs, RBM20 overexpression decreased l-type Ca2+ currents, whereas RBM20 siRNA knockdown increased l-type Ca2+ currents. Finally, we found that RBM20 overexpression reduced CaV1.2 membrane surface expression in NRCMs. Taken together, our results suggest that RBM20 specifically regulates the inclusion of exon 9* in CACNA1C mRNA, resulting in reduced cell-surface membrane expression of l-channels in cardiomyocytes.

Direct, gabapentin-insensitive interaction of a soluble form of the calcium channel subunit \u03b12\u03b4-1 with thrombospondin-4

The α2δ‐1 subunit of voltage-gated calcium channels binds to gabapentin and pregabalin, mediating the analgesic action of these drugs against neuropathic pain. Extracellular matrix proteins from the thrombospondin (TSP) family have been identified as ligands of α2δ‐1 in the CNS. This interaction was found to be crucial for excitatory synaptogenesis and neuronal sensitisation which in turn can be inhibited by gabapentin, suggesting a potential role in the pathogenesis of neuropathic pain. Here, we provide information on the biochemical properties of the direct TSP/α2δ-1 interaction using an ELISA-style ligand binding assay. Our data reveal that full-length pentameric TSP-4, but neither TSP-5/COMP of the pentamer-forming subgroup B nor TSP-2 of the trimer-forming subgroup A directly interact with a soluble variant of α2δ-1 (α2δ-1S). Interestingly, this interaction is not inhibited by gabapentin on a molecular level and is not detectable on the surface of HEK293-EBNA cells over-expressing α2δ‐1 protein. These results provide biochemical evidence that supports a specific role of TSP-4 among the TSPs in mediating the binding to neuronal α2δ‐1 and suggest that gabapentin does not directly target TSP/α2δ-1 interaction to alleviate neuropathic pain.

MiRNA-Coordinated Schizophrenia Risk Networks Cross-Talk with Cardiovascular Repair and Oppose Gliomagenesis

Background: Schizophrenia risk genes are widely investigated, while systemic analysis of miRNA contributing to schizophrenia is lacking. Methods: Schizophrenia-associated genetic loci profile was derived from genome-wide association study (GWAS) of Schizophrenia Working Group of the Psychiatric Genomics Consortium (PGC) dataset. Experimentally confirmed relationships between miRNAs and their target genes were retrieved from miRTarBase. Competitive gene set association analysis for miRNA-target regulations was conducted by the multi-marker analysis of genoMic annotation (MAGMA) and further validated by literature-based functional pathway analysis using Pathway Studio. The associations of the targets of three miRNAs with schizophrenia were further validated using a GWAS of antipsychotic treatment response. Results: Three novel schizophrenia-risk miRNA, namely, miR-208b-3p, miR-208a-3p, and miR-494-5p and their targetomes converge on two calcium voltage-gated channel subunit alpha1 C (CACNA1C) and B-cell lymphoma 2 (Bcl-2), well-known as contributors to schizophrenia. Both miR-208a-3p and miR-208b-3p reduce expression of the RNA-binding protein Quaking (QKI), whose suppression commonly contributes to demyelinating model of schizophrenia and to ischemia/reperfusion injury. Both QKI and hsa-miR-494-5p simultaneously contribute to the risk of schizophrenia and serve as the inhibitors of gliomagenesis. Conclusion: Presented results point at an orchestrating role of miRNAs in pathophysiology of schizophrenia. Cellular effects of risk-associated miRNAs align with the primary etiological hypotheses of schizophrenia. The sharing of regulatory networks schizophrenia and other pathologies may explain higher cardiovascular mortality and lower odds of gliomagenesis previously reported in psychiatric patients. Funding Statement: The study was supported the National Natural Science Foundation of China (No. 81471364) and Primary Research & Development Plan of Jiangsu Province (BE2016630). Declaration of Interests: The authors have nothing to disclose. Ethics Approval Statement: Not applicable. All data were acquired from public databases that do not require an ethical approval.

Increased calcium channel in the lamina propria of aging rat.

The alterations of the extracellular matrix (ECM) in lamina propria of the vocal folds are important changes that are associated with decreased vibrations and increased stiffness in aging vocal fold. The aim of this study was to investigate the differences in gene expression of lamina propria using next generation sequencing (NGS) in young and aging rats and to identify genes that affect aging-related ECM changes for developing novel therapeutic target molecule. Among the 40 genes suggested in the NGS analysis, voltage-gated calcium channels (VGCC) subunit alpha1 S (CACNA1S), VGCC auxiliary subunit beta 1 (CACNB1), and VGCC auxiliary subunit gamma 1 (CACNG1) were increased in the lamina propria of the old rats compared to the young rats. The synthesis of collagen I and III in hVFFs decreased after si-CACNA1S and verapamil treatment. The expression and activity of matrix metalloproteinases (MMP)-1 and -8 were increased in hVFFs after the treatment of verapamil. However, there was no change in the expression of MMP-2 and -9. These results suggest that some calcium channels may be related with the alteration of aging-related ECM in vocal folds. Calcium channel has promising potential as a novel therapeutic target for the remodeling ECM of aging lamina propria.

Ion occupancy of the channel pore is critical for triggering excitation-transcription (ET) coupling

During membrane depolarization the voltage-gated calcium channel (VGCC) activates gene expression in excitable cells by means of a signal-transduction pathway termed excitation transcription (ET) coupling. The L-type calcium channel Cav1.2 can drive nuclear activity by either the ERK-CREB pathway, the Ca2+/calmodulin-dependent protein kinase II (CaMKII) cascade, or via the Ca2+-dependent protein phosphatase calcineurin. The ERK-CREB pathway mediates nuclear activity via a direct interaction of the intracellular β subunit of VGCC with the Ras/GRF1 complex.Here we show that ET coupling in HEK293 cells transfected with wt Cav1.2 or the Timothy mutant Cav1.2G406R is mediated by substituting Ca2+ with the impermeable lanthanum (La3+). In the absence of extracellular Ca2+ or La3+, ET coupling was not triggered. This implies that cation occupancy of the selectivity filter, as opposed to calcium influx, plays an essential role in depolarization triggered signaling to the nucleus.ET coupling triggered by membrane depolarization in Cav1.2 transfected HEK293 cells and neuroendocrine PC12 cells was also supported by substituting Ba2+ for Ca2+ as the charge carrier. Since Ba2+ ions do not bind to calmodulin this implies activation of ET coupling via a Ca2+/calmodulin-independent pathway. Together, these results suggest a model whereby nuclear signaling through the ERK-CREB pathway is driven by voltage-dependent conformational change that requires channel pore occupancy and is Ca2+ influx-independent. This model is also consistent with the previous observation that ET coupling can be driven by the Ca2+-impermeable Cav1.2L745P mutant. Thus, the conversion of synaptic stimuli to transcriptional activation is mediated by the metabotropic function (Ca2+-inflow independent) of Cav1.2, similar to the ion-influx independent depolarization-triggered transmitter release and transcription activation mediated by the NMDA receptors.

Calcium absorption in the fluted giant clam, Tridacna squamosa, may involve a homolog of voltage-gated calcium channel subunit α1 (CACNA1) that has an apical localization and displays light-enhanced protein expression in the ctenidium.

In light, giant clams can increase rates of shell formation and growth due to their symbiotic relationship with phototrophic zooxanthellae residing extracellularly in a tubular system. Light-enhanced shell formation necessitates increase in the uptake of Ca2+ from the ambient seawater and the supply of Ca2+ through the hemolymph to the extrapallial fluid, where calcification occurs. In this study, the complete coding cDNA sequence of a homolog of voltage-gated calcium channel subunit α1 (CACNA1), which is the pore-forming subunit of L-type voltage-gated calcium channels (VGCCs), was obtained from the ctenidium (gill) of the giant clam, Tridacna squamosa. It consisted of 6081bp and encoded a 223kDa polypeptide with 2027 amino acids, which was characterized as the α1D subunit of L-type VGCC. Immunofluorescence microscopy demonstrated that CACNA1 had an apical localization in the epithelial cells of filaments and tertiary water channels in the ctenidium of T. squamosa, indicating that it was well positioned to absorb exogenous Ca2+. Additionally, there was a significant increase in the protein abundance of CACNA1 in the ctenidium of individuals exposed to light for 12h. With more pore-forming CACNA1, there could be an increase in the permeation of exogenous Ca2+ into the ctenidial epithelial cells through the apical membrane. Taken together, these results denote that VGCC could augment exogenous Ca2+ uptake through the ctenidium to support light-enhanced shell formation in T. squamosa. Furthermore, they support the proposition that light-enhanced phenomena in giant clams are attributable primarily to the direct responses of the host's transporters/enzymes to light, in alignment with the symbionts' phototrophic activity.

Scratch Migration Assay and Dorsal Skinfold Chamber for In Vitro and In Vivo Analysis of Wound Healing.

Impaired cutaneous wound healing is a major concern for patients suffering from diabetes and for elderly people, and there is a need for an effective treatment. Appropriate in vitro and in vivo approaches are essential for the identification of new target molecules for drug treatments to improve the skin wound healing process. We identified the β3 subunit of voltage-gated calcium channels (Cavβ3) as a potential target molecule to influence the wound healing in two independent assays, i.e., the in vitro scratch migration assay and the in vivo dorsal skinfold chamber model. Primary mouse embryonic fibroblasts (MEFs) acutely isolated from wild-type (WT) and Cavβ3-deficient mice (Cavβ3 KO) or fibroblasts acutely isolated from WT mice treated with siRNA to down-regulate the expression of the Cacnb3 gene, encoding Cavβ3, were used. A scratch was applied on a confluent cell monolayer and the gap closure was followed by taking microscopic images at defined time points until complete repopulation of the gap by the migrating cells. These images were analyzed, and the cell migration rate was determined for each condition. In an in vivo assay, we implanted a dorsal skinfold chamber on WT and Cavβ3 KO mice, applied a defined circular wound of 2 mm diameter, covered the wound with a glass coverslip to protect it from infections and desiccation, and monitored the macroscopic wound closure over time. Wound closure was significantly faster in Cacnb3-gene-deficient mice. Because the results of the in vivo and the in vitro assays correlate well, the in vitro assay may be useful for the high-throughput screening before validating the in vitro hits by the in vivo wound healing model. What we have shown here for wild-type and Cavβ3-deficient mice or cells might also be applicable for specific molecules other than Cavβ3.

L-type voltage-gated calcium channel regulation of in vitro human cortical neuronal networks

The combination of in vitro multi-electrode arrays (MEAs) and the neuronal differentiation of stem cells offers the capability to study human neuronal networks from patient or engineered human cell lines. Here, we use MEA-based assays to probe synaptic function and network interactions of hiPSC-derived neurons. Neuronal network behaviour first emerges at approximately 30 days of culture and is driven by glutamate neurotransmission. Over a further 30 days, inhibitory GABAergic signalling shapes network behaviour into a synchronous regular pattern of burst firing activity and low activity periods. Gene mutations in L-type voltage gated calcium channel subunit genes are strongly implicated as genetic risk factors for the development of schizophrenia and bipolar disorder. We find that, although basal neuronal firing rate is unaffected, there is a dose-dependent effect of L-type voltage gated calcium channel inhibitors on synchronous firing patterns of our hiPSC-derived neural networks. This demonstrates that MEA assays have sufficient sensitivity to detect changes in patterns of neuronal interaction that may arise from hypo-function of psychiatric risk genes. Our study highlights the utility of in vitro MEA based platforms for the study of hiPSC neural network activity and their potential use in novel compound screening.

Anxiolytic effects of the novel α2δ ligand mirogabalin in a rat model of chronic constriction injury, an experimental model of neuropathic pain.

Psychiatric disorders such as anxiety and depression are frequently observed in neuropathic pain patients, and negatively impact their quality of life. Mirogabalin is a novel ligand for the α2δ subunit of voltage-gated calcium channels and has unique binding characteristics to α2δ subunits and potent and long-lasting analgesic effects in neuropathic pain models. To provide further information on the pharmacological profile of mirogabalin and its utility for chronic pain therapy, we investigated its anxiolytic effects in an experimental animal model for neuropathic pain. In chronic constriction injury (CCI) model rats, mechanical hypersensitivity was determined by the von Frey test. Anxiety- and depression-related behaviours were evaluated using the elevated plus maze test and forced swimming test, respectively. CCI model rats showed sustained tactile allodynia followed by anxiety-related behaviours, not depression-related behaviours. The tactile allodynia (significant decreases in paw withdrawal threshold) developed within 2weeks after model preparation, whereas the anxiety-related behaviours (significant decreases in the number of entries and time spent in open arms and significant increases in time spent in closed arms) were observed at 5weeks but not 4weeks after model preparation. Single oral administration of mirogabalin (3 or 10mg/kg) dose-dependently alleviated the above-mentioned anxiety-related behaviours and tactile allodynia. CCI model rats showed anxiety-related behaviours in a time-dependent manner in the elevated plus maze test. Mirogabalin alleviated both the anxiety-related behaviours and tactile allodynia in CCI model rats. Mirogabalin may provide effective anxiety relief as well as pain relief in patients with neuropathic pain.

Genes Involved in Neurodevelopment, Neuroplasticity and Major Depression: No Association for CACNA1C, CHRNA7 and MAPK1.

ObjectiveGenetics factors are likely to play a role in the risk, clinical presentation and treatment outcome in major depressive disorder (MDD). In this study, we investigated the role of three candidate genes for MDD; calcium voltage-gated channel subunit alpha1 C (CACNA1C ), cholinergic receptor nicotinic alpha 7 subunit (CHRNA7 ), and mitogen-activated protein kinase 1 (MAPK1).MethodsTwo-hundred forty-two MDD patients and 326 healthy controls of Korean ancestry served as samples for the analyses. Thirty-nine single nucleotide polymorphisms (SNPs) within CACNA1C, CHRNA7, and MAPK1 genes were genotyped and subsequently tested for association with MDD (primary analysis) and other clinical features (symptoms’ severity, age of onset, history of suicide attempt, treatment outcome) (secondary analyses). Single SNPs, haplotypes and epistatic analyses were performed.ResultsSingle SNPs were not associated with disease risk and clinical features. However, a combination of alleles (haplotype) within MAPK1 was found associated with MDD-status. Secondary analyses detected a possible involvement of CACNA1C haplotype in resistance to antidepressant treatment.ConclusionThese data suggest a role for MAPK1 and CACNA1C in MDD risk and treatment resistance, respectively. However, since many limitations characterize the analysis, the results must be considered with great caution and verified.

Association of CACNA1C polymorphisms with serum BDNF levels in bipolar disorder.

Variation in the CACNA1C gene has been associated with bipolar disorder in several genome-wide association studies. This gene encodes the alpha 1C subunit of L-type voltage-gated calcium channels, which play an essential role in neurons. We analysed 39 biomarkers in either cerebrospinal fluid or serum in relation to six different CACNA1C variants in 282 patients with bipolar disorder and 90 controls. We report associations of CACNA1C risk alleles with serum levels of BDNF as well as tissue plasminogen activator, which converts pro-BDNF to mature BDNF. This sheds light on links between CACNA1C genetic variants and pathophysiological mechanisms in bipolar disorder.

Neuropeptide FF (FLFQPQRF-NH2) and its Fragments Bind to α2δ Subunit of Voltage-Gated Calcium Channels.

Gabapentin, a drug for neuropathic pain, exerts its therapeutic effect via inhibition of the a2d subunit of N-type Ca2+ channels. Thus, finding peptides that specifically displace gabapentin from its binding site may lead to the development of new drugs. Displacement of bound [3H]-gabapentin in membrane preparations of rat cerebral cortex and of human Cav2.2/β3/α2δ1 expressed in CHO cell line. Neuropeptide FLFQPQRF-NH2 specifically displaced bound [3H]-gabapentin in membrane preparations from rats and CHO cells. Truncation of the C-terminus of FLFQPQRF-NH2 by three amino acid residues to produce FLFQP-NH2 improved the displacement of gabapentin. FLFQP-NH2 displaced bound [3H]-gabapentin with IC50 and Ki values of 2.7 µM and 1.7 µM, respectively. Deletion of two amino acid residues (FQ) in the middle of the FLFQP-NH2 sequence yielded FLP-NH2 that displaced bound [3H]-gabapentin with a lower affinity. IC50 and Ki values were 11.9 µM and 7.8 µM, respectively. Neutral binding cooperativity existed when of FLFQP-NH2, FLP- NH2 and gabapentin when incubated together. FLFQPQRF-NH2 but not FLFQP-NH2 displaced bound [3H]- gabapentin to membrane preparations of human Cav2.2/b3/a2d1 expressed in CHO cells. FLFQPQRF-NH2, FLFQP-NH2 and FLP-NH2 displace bound gabapentin in membrane preparations of rat cerebral cortex. Binding cooperativity was detected when GBP/FLFQP-NH2/FLP-NH2 were incubated together. These novel binding sites may provide new approaches to modulate L-type Ca2+ channels.

Detection of copy number variation and selection signatures on the X chromosome in Chinese indigenous sheep with different types of tail.

ObjectiveChinese indigenous sheep breeds can be classified into the following three categories by their tail morphology: fat-tailed, fat-rumped and thin-tailed sheep. The typical sheep breeds corresponding to fat-tailed, fat-rumped, and thin-tailed sheep are large-tailed Han, Altay, and Tibetan sheep, respectively. Detection of copy number variation (CNV) and selection signatures provides information on the genetic mechanisms underlying the phenotypic differences of the different sheep types.MethodsIn this study, PennCNV software and F-statistics (FST) were implemented to detect CNV and selection signatures, respectively, on the X chromosome in three Chinese indigenous sheep breeds using ovine high-density 600K single nucleotide polymorphism arrays.ResultsIn large-tailed Han, Altay, and Tibetan sheep, respectively, a total of six, four and 22 CNV regions (CNVRs) with lengths of 1.23, 0.93, and 7.02 Mb were identified on the X chromosome. In addition, 49, 34, and 55 candidate selection regions with respective lengths of 27.49, 16.47, and 25.42 Mb were identified in large-tailed Han, Altay, and Tibetan sheep, respectively. The bioinformatics analysis results indicated several genes in these regions were associated with fat, including dehydrogenase/reductase X-linked, calcium voltage-gated channel subunit alpha1 F, and patatin like phospholipase domain containing 4. In addition, three other genes were identified from this analysis: the family with sequence similarity 58 member A gene was associated with energy metabolism, the serine/arginine-rich protein specific kinase 3 gene was associated with skeletal muscle development, and the interleukin 2 receptor subunit gamma gene was associated with the immune system.ConclusionThe results of this study indicated CNVRs and selection regions on the X chromosome of Chinese indigenous sheep contained several genes associated with various heritable traits.

Discovery of Biomarker Panels for Neural Dysfunction in Inborn Errors of Amino Acid Metabolism

Patients with inborn errors of amino acid metabolism frequently show neuropsychiatric symptoms despite accurate metabolic control. This study aimed to gain insight into the underlying mechanisms of neural dysfunction. Here we analyzed the expression of brain-derived neurotrophic factor (BDNF) and 10 genes required for correct brain functioning in plasma and blood of patients with Urea Cycle Disorders (UCD), Maple Syrup Urine Disease (MSUD) and controls. Receiver-operating characteristic (ROC) analysis was used to evaluate sensitivity and specificity of potential biomarkers. CACNA2D2 (α2δ2 subunit of voltage-gated calcium channels) and MECP2 (methyl-CpG binding protein 2) mRNA and protein showed an excellent neural function biomarker signature (AUC ≥ 0,925) for recognition of MSUD. THBS3 (thrombospondin 3) mRNA and AABA gave a very good biomarker signature (AUC 0,911) for executive-attention deficits. THBS3, LIN28A mRNA, and alanine showed a perfect biomarker signature (AUC 1) for behavioral and mood disorders. Finally, a panel of BDNF protein and at least two large neural AAs showed a perfect biomarker signature (AUC 1) for recognition of psychomotor delay, pointing to excessive protein restriction as central causative of psychomotor delay. To conclude, our study has identified promising biomarker panels for neural function evaluation, providing a base for future studies with larger samples.

Proestrus Differentially Regulates Expression of Ion Channel and Calcium Homeostasis Genes in GnRH Neurons of Mice.

In proestrus, the changing gonadal hormone milieu alters the physiological properties of GnRH neurons and contributes to the development of the GnRH surge. We hypothesized that proestrus also influences the expression of different ion channel genes in mouse GnRH neurons. Therefore, we performed gene expression profiling of GnRH neurons collected from intact, proestrous and metestrous GnRH-GFP transgenic mice, respectively. Proestrus changed the expression of 37 ion channel and 8 calcium homeostasis-regulating genes. Voltage-gated sodium channels responded with upregulation of three alpha subunits (Scn2a1, Scn3a, and Scn9a). Within the voltage-gated potassium channel class, Kcna1, Kcnd3, Kcnh3, and Kcnq2 were upregulated, while others (Kcna4, Kcnc3, Kcnd2, and Kcng1) underwent downregulation. Proestrus also had impact on inwardly rectifying potassium channel subunits manifested in enhanced expression of Kcnj9 and Kcnj10 genes, whereas Kcnj1, Kcnj11, and Kcnj12 subunit genes were downregulated. The two-pore domain potassium channels also showed differential expression with upregulation of Kcnk1 and reduced expression of three subunit genes (Kcnk7, Kcnk12, and Kcnk16). Changes in expression of chloride channels involved both the voltage-gated (Clcn3 and Clcn6) and the intracellular (Clic1) subtypes. Regarding the pore-forming alpha-1 subunits of voltage-gated calcium channels, two (Cacna1b and Cacna1h) were upregulated, while Cacna1g showed downregulation. The ancillary subunits were also differentially regulated (Cacna2d1, Cacna2d2, Cacnb1, Cacnb3, Cacnb4, Cacng5, Cacng6, and Cacng8). In addition, ryanodine receptor 1 (Ryr1) gene was downregulated, while a transient receptor potential cation channel (Trpm3) gene showed enhanced expression. Genes encoding proteins regulating the intracellular calcium homeostasis were also influenced (Calb1, Hpca, Hpcal1, Hpcal4, Cabp7, Cab 39l, and Cib2). The differential expression of genes coding for ion channel proteins in GnRH neurons at late proestrus indicates that the altering hormone milieu contributes to remodeling of different kinds of ion channels of GnRH neurons, which might be a prerequisite of enhanced cellular activity of GnRH neurons and the subsequent surge release of the neurohormone.

Combined effects of the rs9810888 polymorphism in calcium voltage-gated channel subunit alpha1 D (CACNA1D) and lifestyle behaviors on blood pressure level among Chinese children.

BackgroundsA recent GWAS Study found a new locus (rs9810888 in CACNA1D) was associated with blood pressure (BP) in Chinese adults. But whether the association exists in children is unknown. Whether lifestyle behaviors could interact with rs9810888 on BP is not clear. This study aimed to identify the association between rs9810888 and BP in children, and also explore the gene-lifestyle interaction.MethodsA case-control study was conducted among 2030 Chinese children aged 7 to 18 years. Genotyping was conducted by using the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Lifestyle behaviors were investigated with questionnaire.ResultsWith adjustment for age, age square, sex, study group and body mass index (BMI), rs9810888 was significantly associated with diastolic BP (DBP) (b = 1.69, p = 0.021) and mean arterial BP (MAP) (b = 1.56, p = 0.010). Stratified analysis showed that the rs9810888 GG genotype carriers had higher DBP than GT/TT carriers (b = 3.78, p = 0.023) in the subgroup having protein intake (meat/fish/soybeans/egg) <twice/day, but not in the subgroup ≥twice/day. In addition, in the subgroup with screen time≥2h/day, the rs9810888 GG genotype carriers had higher DBP than GT/TT carriers (b = 5.49, p = 0.012), but not in the subgroup with screen time<2h/day. For MAP, in the subgroup having either fruits or vegetables < twice/day, the rs9810888 GG genotype carriers had higher MAP than GT/TT carriers (b = 2.64, p = 0.037), but not in the subgroup with both fruits and vegetables intake ≥twice/day. Additionally, in the subgroup with screen time≥2h/day, the rs9810888 GG genotype carriers had higher MAP than GT/TT carriers (b = 4.80, p = 0.017), but not in those with screen time<2h/day.ConclusionsThe CACNA1D rs9810888 polymorphism was significantly associated with DBP and MAP. In addition, unhealthy lifestyle behaviors and rs9810888 GG genotype had combined effects on BP level among Chinese children.

Six-mRNA risk score system and nomogram constructed for patients with ovarian cancer.

Platinum is a commonly used drug for the treatment of ovarian cancer (OC). The aim of the current study was to design and construct a risk score system for predicting the prognosis of patients with OC receiving platinum chemotherapy. The mRNA sequencing data and copy number variation (CNV) information (training set) of patients with OC were downloaded from The Cancer Genome Atlas database. A validation set, GSE63885, was obtained from Gene Expression Omnibus database. The differentially expressed genes (DEGs) and CNV genes (DECNs) between platinum-resistant and platinum-sensitive groups were identified using the limma package. The intersection between DEGs and DECNs were selected. Cox regression analysis was used to identify the genes and clinical factors associated with prognosis. Risk score system assessment and nomogram analysis were performed using the survival and rms packages in R. Gene Set Enrichment Analysis was used to identify the enriched pathways in high and low risk score groups. From 1,144 DEGs and 1,864 DECNs, 48 genes that occurred in the two datasets were selected. A total of six independent prognostic genes (T-box transcription factor T, synemin, tektin 5, growth differentiation factor 3, solute carrier family 22 member 3 and calcium voltage-gated channel subunit α1 C) and platinum response status were revealed to be associated with prognosis. Based on the six independent prognostic genes, a risk score system was constructed and assessed. Nomogram analysis revealed that the patients with the sensitive status and low risk scores had an improved prognosis. Furthermore, the current study revealed that the 574 DEGs identified were involved in eight pathways, including chemokine signaling pathway, toll-like receptor signaling pathway, cytokine-cytokine receptor interaction, RIG I like receptor signaling pathway, natural killer cell mediated cytotoxicity, apoptosis, T cell receptor signaling pathway and Fc ε receptor 1 signaling pathway. The six-mRNA risk score system designed in the present study may be used as prognosis predictor in patients with OC, whereas the nomogram may be valuable for identifying patients with OC who may benefit from platinum chemotherapy.

CACNA1C (rs1006737) may be a susceptibility gene for schizophrenia: An updated meta-analysis.

IntroductionSchizophrenia is a serious mental illness with a genetic predisposition. Genome‐wide association studies (GWAS) have identified the α‐1C subunit of the L‐type voltage‐gated calcium channel (CACNA1C) gene as a significant risk gene for schizophrenia. However, there are inconsistent conclusions in case–control studies.MethodsWe performed a comprehensive meta‐analysis of all available samples from existing studies under four different genetic models (recessive model, dominant model, additive model and allele model) to further confirm whether CACNA1C rs1006737 is an authentic risk single nucleotide polymorphism (SNP) for schizophrenia.ResultsA statistically significant difference under the four models (all p < 0.05) was observed by pooling nine Asian and European studies, including a total of 12,744 cases and 16,460 controls. For European‐decent samples, a significant difference was identified between patients and controls for the four models (all p < 0.05). We observed a significant difference between patients and controls for the recessive model and allele model (GG vs. GA + AA: p < 0.00001; G vs. A: p < 0.00001) using a fixed effect model, but the dominant model (GG + GA vs. AA: OR: p = 0.15) and additive model (GG vs. AA: p = 0.11) showed no significant difference between patients and controls in the Asian samples.ConclusionOur findings provide important evidence for the establishment of CACNA1C as a susceptibility gene for schizophrenia across world populations, but its roles in the pathogenesis of schizophrenia need to be further investigated.