Deletion of TRPV3 and CaV3.2 T-type channels in mice undermines fertility and Ca2+ homeostasis in oocytes and eggs.
Ontology highlight
ABSTRACT:
SUBMITTER: Mehregan A
PROVIDER: S-EPMC8313860 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:STUDY QUESTION:Are the transient receptor potential cation channels vanilloid 3 (TRPV3) present and able to mediate strontium (Sr2+) induced artificial activation in human oocytes? SUMMARY ANSWER:Sr2+ did not induce Ca2+ rises or provoke activation in human oocytes, however, mRNA for the TRPV3 channel was present in metaphase II (MII) human oocytes after IVM and TRPV3 agonists induced Ca2+ rises and oocyte activation, demonstrating the channels were functional. WHAT IS KNOWN ALREADY:Selective activation of TRPV3 by agonists induces Ca2+ entry and promotes mouse oocyte activation, and the absence of TRPV3 channels in mouse oocytes prevents Sr2+ mediated artificial activation. Sr2+ is sometimes used to overcome fertilization failure after ICSI in the clinic, but its efficiency is still controversial and the mechanism(s) of how it mediates the Ca2+ flux has not been studied yet in human. STUDY DESIGN SIZE DURATION:The protein distribution (n = 10) and mRNA expression level (n = 19) of the TRPV3 channels was investigated in human MII oocytes after IVM. The Sr2+ (10 mM) and TRPV3 agonists (200 ?M 2-aminoethoxydiphenyl borate [2-APB] and 200 ?M carvacrol)-induced Ca2+ response was analyzed in human (n = 15, n = 16 and n = 16, respectively) and mouse oocytes (n = 15, n = 19 and n = 26, respectively). The subsequent embryonic developmental potential following the parthenogenetic activation using these three agents was recorded in human (n = 10, n = 9 and n = 9, respectively) and mouse (n = 20 per agent) oocytes, by determining pronucleus, or 2-cell and blastocyst formation rates. PARTICIPANTS/MATERIALS SETTING METHODS:MII oocytes from B6D2F1 mice (6-10 weeks old) as well as human IVM oocytes and IVO oocytes (from patients aged 25-38 years old) with aggregates of smooth endoplasmic reticulum clusters were used. The expression of TRPV3 channels was determined by immunofluorescence staining with confocal microscopy and RT-PCR, and the temporal evolution of intracellular Ca2+ concentration was measured by time-lapse imaging after exposure to Sr2+ and TRPV3 agonists (2-APB and carvacrol). Artificial activation efficiency was assessed using these agents. MAIN RESULTS AND THE ROLE OF CHANCE:Sr2+ did not promote Ca2+ oscillations or provoke activation in human oocytes. Transcripts of TRPV3 channels were present in IVM MII human oocytes. TRPV3 protein was expressed and distributed throughout the ooplasm of human oocytes, rather than particularly concentrated in plasma membrane as observed in mouse MII oocytes. Both agonists of TRPV3 (2-APB and carvacrol), promoted a single Ca2+ transient and activated a comparable percentage of more than half of the exposed human oocytes (P > 0.05). The agonist 2-APB was also efficient in activating mouse oocytes, however, significantly fewer mouse oocytes responded to carvacrol than 2-APB in both the Ca2+ analysis and activation test (P < 0.001). LIMITATIONS REASONS FOR CAUTION:The availability of fresh IVO matured oocytes in human was limited. Data from TRPV3 knockout model are not included. WIDER IMPLICATIONS OF THE FINDINGS:The benefit of clinical application using Sr2+ to overcome fertilization failure after ICSI requires further validation. STUDY FUNDING/COMPETING INTERESTS:This study was supported by FWO-Vlaanderen, China Scholarship Council and Special Research Fund from Ghent University (Bijzonder Onderzoeksfonds, BOF). No competing interests are declared.
| S-EPMC6276696 | biostudies-literature
Project description:Functional modifications of neuronal P/Q-type voltage-dependent Ca2+ channels expressed in Xenopus oocytes by oxidation were examined electrophysiologically. Oxidation by external H2O2 enhanced the whole-oocyte currents through the Ca2+ channels composed of the alpha1A, alpha2/delta, and beta3 subunits at negative voltages (<0 mV) without markedly affecting the currents at more positive voltages. Single-channel analysis showed that oxidation accelerates the overall channel opening process. The effect of H2O2 to enhance the Ca2+ channel activity did not require heterologous expression of the alpha2/delta subunit, and it was not mimicked by a cysteine-specific oxidizing agent. The results suggest that oxidative stress may regulate the activity of neuronal Ca2+ channels and that regulation by oxidation may be important in some clinical situations, such as in reperfusion injury after ischemic episodes.
| S-EPMC6792958 | biostudies-literature
Project description:In briefOocyte quality remains the most important and unsolved issue in reproduction. Our data show that multidrug resistance transporters and oocyte mitochondria are involved in determining oocyte quality in a mouse model.AbstractMultidrug resistance transporter-1 (MDR-1) is a transmembrane ATP-dependent effluxer present in organs that transport a variety of xenobiotics and by-products. Previous findings by our group demonstrated that this transporter is also present in the oocyte mitochondrial membrane and that its mutation led to abnormal mitochondrial homeostasis. Considering the importance of these organelles in the female gamete, we assessed the impact of MDR-1 dysfunction on mouse oocyte quality, with a particular focus on the meiotic spindle organization, aneuploidies, Ca2+ homeostasis, ATP production and mtDNA mutations. Our results demonstrate that young Mdr1a mutant mice produce oocytes characterized by lower quality, with a significant delay in the germinal vesicle to germinal vesicle breakdown transition, an increased percentage of symmetric divisions, chromosome misalignments and a severely altered meiotic spindle shape compared to the wild types. Mutant oocytes exhibit 7000 more SNPs in the exomic DNA and twice the amount of mitochondrial DNA (mtDNA) SNPs compared to the wild-type ones. Ca2+ analysis revealed the inability of MDR-1 mutant oocytes to manage Ca2+ storage content and oscillations in response to several stimuli, and ATP quantification shows that mutant oocytes trend toward lower ATP levels compared to wild types. Finally, 1-year-old mutant ovaries express a lower amount of SIRT1, SIRT3, SIRT5, SIRT6 and SIRT7 compared to wild-type levels. These results together emphasize the importance of MDR-1 in mitochondrial physiology and highlight the influence of MDR-1 on oocyte quality and ovarian aging.
| S-EPMC9782432 | biostudies-literature
Project description:Chronic intermittent hypoxia (CIH) is a hallmark manifestation of sleep apnea. A heightened carotid body activity and the resulting chemosensory reflex mediate increased sympathetic nerve activity by CIH. However, the mechanisms underlying heightened carotid body activity by CIH are not known. An elevation of intracellular calcium ion concentration ([Ca(2+)]i) in glomus cells, the primary oxygen-sensing cells, is an essential step for carotid body activation by hypoxia. In the present study, we examined the effects of CIH on the glomus cell [Ca(2+)]i response to hypoxia and assessed the underlying mechanisms. Glomus cells were harvested from adult rats or wild-type mice treated with 10 days of either room air (control) or CIH (alternating cycles of 15 s of hypoxia and 5 min of room air; 9 episodes/h; 8 h/day). CIH-treated glomus cells exhibited an enhanced [Ca(2+)]i response to hypoxia, and this effect was absent in the presence of 2-(4-cyclopropylphenyl)-N-((1R)-1-[5-[(2,2,2-trifluoroethyl)oxo]-pyridin-2-yl]ethyl)acetamide (TTA-A2), a specific inhibitor of T-type Ca(2+) channels, and in voltage-gated calcium channel, type 3.2 (CaV3.2), null glomus cells. CaV3.2 knockout mice exhibited an absence of CIH-induced hypersensitivity of the carotid body. CIH increased reactive oxygen species (ROS) levels in glomus cells. A ROS scavenger prevented the exaggerated TTA-A2-sensitive [Ca(2+)]i response to hypoxia. CIH had no effect on CaV3.2 mRNA levels. CIH augmented Ca(2+) currents and increased CaV3.2 protein in plasma membrane fractions of human embryonic kidney-293 cells stably expressing CaV3.2, and either a ROS scavenger or brefeldin-A, an inhibitor of protein trafficking, prevented these effects. These findings suggest that CIH leads to an augmented Ca(2+) influx via ROS-dependent facilitation of CaV3.2 protein trafficking to the plasma membrane.
| S-EPMC4760493 | biostudies-literature
Project description:In mammals, calcium influx is required for oocyte maturation and egg activation. The molecular identities of the calcium-permeant channels that underlie the initiation of embryonic development are not established. Here, we describe a transient receptor potential (TRP) ion channel current activated by TRP agonists that is absent in TrpV3(-/-) eggs. TRPV3 current is differentially expressed during oocyte maturation, reaching a peak of maximum density and activity at metaphase of meiosis II (MII), the stage of fertilization. Selective activation of TRPV3 channels provokes egg activation by mediating massive calcium entry. Widely used to activate eggs, strontium application is known to yield normal offspring in combination with somatic cell nuclear transfer. We show that TRPV3 is required for strontium influx, because TrpV3(-/-) eggs failed to conduct Sr(2+) or undergo strontium-induced activation. We propose that TRPV3 is a major mediator of calcium influx in mouse eggs and is a putative target for artificial egg activation.
| S-EPMC3918412 | biostudies-literature
Project description:The present study was undertaken to explore the relative contributions of Cav3.2 T-type channels to mediating the antihyperalgesic activity of joint manipulation (JM) therapy. We used the chronic constriction injury model (CCI) to induce peripheral neuropathy and chronic pain in male mice, followed by JM. We demonstrate that JM produces long-lasting mechanical anti-hyperalgesia that is abolished in Cav3.2 null mice. Moreover, we found that JM displays a similar analgesic profile as the fatty acid amide hydrolase inhibitor URB597, suggesting a possible converging mechanism of action involving endocannabinoids. Overall, our findings advance our understanding of the mechanisms through which JM produces analgesia.
| S-EPMC10355051 | biostudies-literature
Project description:Cav3 channels play an important role in modulating chronic pain. However, less is known about the functional changes of Cav3 channels in superficial spinal dorsal horn in neuropathic pain states. Here, we examined the effect of partial sciatic nerve ligation (PSNL) on either expression or electrophysiological properties of Cav3 channels in superficial spinal dorsal horn. Our in vivo studies showed that the blockers of Cav3 channels robustly alleviated PSNL-induced mechanical allodynia and thermal hyperalgesia, which lasted at least 14 days following PSNL. Meanwhile, PSNL triggered an increase in both mRNA and protein levels of Cav3.2 but not Cav3.1 or Cav3.3 in rats. However, in Cav3.2 knockout mice, PSNL predominantly attenuated mechanical allodynia but not thermal hyperalgesia. In addition, the results of whole-cell patch-clamp recordings showed that both the overall proportion of Cav3 current-expressing neurons and the Cav3 current density in individual neurons were elevated in spinal lamina II neurons from PSNL rats, which could not be recapitulated in Cav3.2 knockout mice. Altogether, our findings reveal that the elevated functional Cav3.2 channels in superficial spinal dorsal horn may contribute to the mechanical allodynia in PSNL-induced neuropathic pain model.
| S-EPMC6458665 | biostudies-literature
Project description:In starfish, the addition of the hormone 1-methyladenine (1-MA) to immature oocytes (germinal vesicle, GV-stage) arrested at the prophase of the first meiotic division induces meiosis resumption (maturation), which makes the mature eggs able to respond to the sperm with a normal fertilization response. The optimal fertilizability achieved during the maturation process results from the exquisite structural reorganization of the actin cytoskeleton in the cortex and cytoplasm induced by the maturing hormone. In this report, we have investigated the influence of acidic and alkaline seawater on the structure of the cortical F-actin network of immature oocytes of the starfish (Astropecten aranciacus) and its dynamic changes upon insemination. The results have shown that the altered seawater pH strongly affected the sperm-induced Ca2+ response and the polyspermy rate. When immature starfish oocytes were stimulated with 1-MA in acidic or alkaline seawater, the maturation process displayed a strong dependency on pH in terms of the dynamic structural changes of the cortical F-actin. The resulting alteration of the actin cytoskeleton, in turn, affected the pattern of Ca2+ signals at fertilization and sperm penetration.
| S-EPMC10000582 | biostudies-literature
Project description:The hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are subthreshold, voltage-gated ion channels that are highly expressed in hippocampal and cortical pyramidal cell dendrites, where they are important for regulating synaptic potential integration and plasticity. We found that HCN1 subunits are also localized to the active zone of mature asymmetric synaptic terminals targeting mouse entorhinal cortical layer III pyramidal neurons. HCN channels inhibited glutamate synaptic release by suppressing the activity of low-threshold voltage-gated T-type (Ca(V)3.2) Ca²(+) channels. Consistent with this, electron microscopy revealed colocalization of presynaptic HCN1 and Ca(V)3.2 subunit. This represents a previously unknown mechanism by which HCN channels regulate synaptic strength and thereby neural information processing and network excitability.
| S-EPMC3068302 | biostudies-literature
Project description:BackgroundChromodomain helicase DNA-binding protein 7 (CHD7), which is associated with CHARGE (Coloboma, Heart defect, Atresia choanae, Restricted growth, Genital hypoplasia and Ear abnormality) syndrome is an important regulator in many vital developmental processes. However, its role during oocyte development remains unknown.MethodsWe screened the Gene Expression Omnibus (GEO) database for expression levels of CHD7 during folliculogenesis. We generated a conditional knockout (cKO) mouse strain with oocyte-specific deletion of CHD7 (Gdf9-Cre:Chd7f/f ) using the Cre-loxP approach. Evaluation of follicle numbers and reproductive ability was then conducted. In addition, granulosa cell (GC) apoptosis was assessed by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay and cleaved caspase-3, using immunohistochemistry (IHC) and immunofluorescence (IF). GC proliferation was measured by Ki67 staining as evaluated by IHC.ResultsIn our study, we demonstrated that CHD7 has high expression throughout all developmental stages of the oocyte. We found that deletion of Chd7 in oocytes can cause infertility or sub-fertility in female mice and is associated with decreased follicle numbers at all stages. In addition, we found that GC apoptosis was significantly higher in cKO mice.ConclusionsTo our knowledge, our study has been the first to show that CHD7 plays a specific role during oogenesis. Our findings provide new insights into CHD7-related infertility.
| S-EPMC8987892 | biostudies-literature