Kisspeptin-10 Facilitates a Plasma Membrane-Driven Calcium Oscillator in Gonadotropin-Releasing Hormone-1 Neurons

Kisspeptins, the natural ligands of the G-protein-coupled receptor (GPR)-54, are the most potent stimulators of GnRH-1 secretion and as such are critical to reproductive function. However, the mechanism by which kisspeptins enhance calcium-regulated neuropeptide secretion is not clear. In the present study, we used GnRH-1 neurons maintained in mice nasal explants to examine the expression and signaling of GPR54. Under basal conditions, GnRH-1 cells exhibited spontaneous baseline oscillations in intracellular calcium concentration ([Ca2+]i), which were critically dependent on the operation of voltage-gated, tetrodotoxin (TTX)-sensitive sodium channels and were not coupled to calcium release from intracellular pools. Activation of native GPR54 by kisspeptin-10 initiated [Ca2+]i oscillations in quiescent GnRH-1 cells, increased the frequency of calcium spiking in oscillating cells that led to summation of individual spikes into plateau-bursting type of calcium signals in a subset of active cells. These changes predominantly reflected the stimulatory effect of GPR54 activation on the plasma membrane oscillator activity via coupling of this receptor to phospholipase C signaling pathways. Both components of this pathway, inositol 1,3,4-trisphosphate and protein kinase C, contributed to the receptor-mediated modulation of baseline [Ca2+]i oscillations. TTX and 2-aminoethyl diphenylborinate together abolished agonist-induced elevation in [Ca2+]i in almost all cells, whereas flufenamic acid was less effective. Together these results indicate that a plasma membrane calcium oscillator is spontaneously operative in the majority of prenatal GnRH-1 neurons and is facilitated by kisspeptin-10 through phosphatidyl inositol diphosphate hydrolysis and depolarization of neurons by activating TTX-sensitive sodium channels and nonselective cationic channels. GnRH-1 neurons exhibit a spontaneously active calcium oscillator, dependent on tetrodotoxin-sensitive sodium conductance. Kisspeptin-10/GPR54, via phosphatidyl inositol diphosphate 2 hydrolysis, utilizes these channels and non-selective cationic channels.

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Chromosomal instability in different species of agricultural animals (review)

Glavnyj zootehnik (Head of Animal Breeding) ◽

10.33920/sel-03-2105-03 ◽

2021 ◽

pp. 19-26

Author(s):

V. A. Andreeva

Keyword(s):

Chromosomal Instability ◽

Animal Species ◽

Western Siberia ◽

Reproductive Function ◽

Farm Animals ◽

Chromosomal Disorders ◽

Important Indicator ◽

Karyotypic Analysis ◽

Different Types ◽

Almost All

The purpose of the work was to analyze the frequency of chromosomal instability in different species of agricultural animals in Western Siberia on the basis of literature data. The analysis of the literature on the topic of somatic chromosomal instability in agricultural animals has been carried out. Despite the stability of the chromosome set, deviations from it are quite common. It is noteworthy that chromosomal instability is characteristic of almost all individuals in the population and serves as an important indicator for assessing the natural mutability of chromosomes. It has been found if an aberration appeared in one tissue, it is very likely to occur in others. It has been noted that somatic chromosomal instability occurs in animals with reduced reproductive function, as well as those suffering from any pathology. For example, in calves with parakeratosis the increase in the frequency of chromatid and isochromatid breaks has been found. Analysis of the frequency of aberrations in sires, which differ in the level of perinatal mortality of offspring revealed the increased percentage of offspring mortality in fathers with the large number of chromosomal disorders. Therefore, low fertilization, spontaneous abortions and stillbirths may be indications for karyotypic analysis. It has been revealed that the lability of the karyotype is inherent in all animal species, regardless of the species, sex and age, as well as the morphofunctional state. The data on some types of somatic chromosomal instability in different types of farm animals in Western Siberia has been presented. Such indicators as the frequency of polyploidy, the number of cells with fragments of chromosomes, as well as single and paired fragments of chromosomes has been given. Different types of chromosomal instability varied depending on the animal species and climate zone. The presented data can be accepted as a physiological norm and used in veterinary medicine and animal science.

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Resistance to miltefosine results from amplification of the RTA3 floppase or inactivation of flippases in Candida parapsilosis

10.1101/2021.12.16.473093 ◽

2021 ◽

Author(s):

Sean Bergin ◽

Fang Zhao ◽

Adam P Ryan ◽

Carolin A Müller ◽

Conrad A Nieduszynski ◽

...

Keyword(s):

Plasma Membrane ◽

Copy Number ◽

Candida Parapsilosis ◽

Copy Number Variations ◽

Loss Of Function ◽

Adaptive Laboratory Evolution ◽

Pathogenic Yeast ◽

Lipid Asymmetry ◽

Evolution Experiment ◽

Almost All

Flippases and floppases are two classes of proteins that have opposing functions in the maintenance of lipid asymmetry of the plasma membrane. Flippases translocate lipids from the exoplasmic leaflet to the cytosolic leaflet, and floppases act in the opposite direction. Phosphatidylcholine (PC) is a major component of the eukaryotic plasma membrane and is asymmetrically distributed, being more abundant in the exoplasmic leaflet. Here we show that gene amplification of a putative PC floppase or double disruption of two PC flippases in the pathogenic yeast Candida parapsilosis results in resistance to miltefosine, an alkylphosphocholine drug that affects PC metabolism that has recently been granted orphan drug designation approval by the US FDA for treatment of invasive candidiasis. We analysed the genomes of 170 C. parapsilosis isolates and found that 107 of them have copy number variations (CNVs) at the RTA3 gene. RTA3 encodes a putative PC floppase whose deletion is known to increase the inward translocation of PC in Candida albicans. RTA3 copy number ranges from 2 to >40 across the C. parapsilosis isolates. Interestingly, 16 distinct CNVs with unique endpoints were identified, and phylogenetic analysis shows that almost all of them have originated only once. We found that increased copy number of RTA3 correlates with miltefosine resistance. Additionally, we conducted an adaptive laboratory evolution experiment in which two C. parapsilosis isolates were cultured in increasing concentrations of miltefosine over 26 days. Two genes, CPAR2_303950 and CPAR2_102700, gained homozygous protein-disrupting mutations in the evolved strains and code for putative PC flippases homologous to S. cerevisiae DNF1. Our results indicate that alteration of lipid asymmetry across the plasma membrane is a key mechanism of miltefosine resistance. We also find that C. parapsilosis is likely to gain resistance to miltefosine rapidly, because many isolates carry loss-of-function alleles in one of the flippase genes.

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Calcium-induced calcium release mechanism in guinea pig taenia caeci.

The Journal of General Physiology ◽

10.1085/jgp.94.2.363 ◽

1989 ◽

Vol 94 (2) ◽

pp. 363-383 ◽

Cited By ~ 142

Author(s):

M Iino

Keyword(s):

Guinea Pig ◽

Calcium Release ◽

Adenine Nucleotide ◽

Physiological Role ◽

Release Mechanism ◽

Smooth Muscle Fiber ◽

Skinned Smooth Muscle ◽

Intracellular Ca ◽

Almost All ◽

Calcium Induced Calcium Release

Fura-2 was used to measure the amount of Ca released from the intracellular Ca store of a saponin-skinned smooth muscle fiber bundle of the guinea pig taenia caeci (width, 150-250 microns) placed in a capillary cuvette at 20-22 degrees C. The amount of Ca actively loaded into the store was assayed when released by the application of 50 mM caffeine and/or 10 microM inositol 1,4,5-trisphosphate (IP3) in the absence of ATP, and was found to have a biphasic dependence on the loading [Ca2+] with a peak near pCa 6. After Ca loading at pCa 6, IP3 released almost all the releasable Ca, whereas caffeine discharged Ca from only approximately 40% of the store. The maximum amount of Ca in the store was some 220 mumol/liter cell water. Ca in the caffeine-releasable store was released approximately exponentially to zero with time when Ca2+ was applied in the absence of ATP, and the rate constant of the Ca-induced Ca release (CICR) increased steeply with the concentration of Ca2+ applied. Increase in [Mg2+] (0.5-5.0 mM) or decrease in pH (7.3-6.7) shifted the relation between pCa and the rate of CICR roughly in parallel toward the lower pCa. An adenine nucleotide increased the rate of the CICR, but it did not change the range of effective [Ca2+]. 5 mM caffeine greatly enhanced the CICR mechanism, making it approximately 30 times more sensitive to [Ca2+]. However the drug had no Ca-releasing action in the absence of Ca2+. Procaine in millimolar concentrations inhibited the rate of the CICR. These properties are similar to those of the skeletal muscle CICR and ryanodine receptor channels. Rates of the CICR under a physiological ionic milieu were estimated from the results, and a [Ca2+] greater than 1 microM was expected to be necessary for the activation of the Ca release. This Ca sensitivity seems too low for the CICR mechanism to play a primary physiological role in Ca mobilization, unless assisted by other mechanisms.

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Estradiol activates epithelial sodium channels in rat alveolar cells through the G protein-coupled estrogen receptor

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00008.2013 ◽

2013 ◽

Vol 305 (11) ◽

pp. L878-L889 ◽

Cited By ~ 26

Author(s):

Megan M. Greenlee ◽

Jeremiah D. Mitzelfelt ◽

Ling Yu ◽

Qiang Yue ◽

Billie Jeanne Duke ◽

...

Keyword(s):

Estrogen Receptor ◽

Plasma Membrane ◽

G Protein ◽

Sodium Channels ◽

Female Rats ◽

Channel Activity ◽

Alveolar Cells ◽

Α Subunit ◽

Female Sex ◽

G Protein Coupled

Female sex predisposes individuals to poorer outcomes during respiratory disorders like cystic fibrosis and influenza-associated pneumonia. A common link between these disorders is dysregulation of alveolar fluid clearance via disruption of epithelial sodium channel (ENaC) activity. Recent evidence suggests that female sex hormones directly regulate expression and activity of alveolar ENaC. In our study, we identified the mechanism by which estradiol (E2) or progesterone (P4) independently regulates alveolar ENaC. Using cell-attached patch clamp, we measured ENaC single-channel activity in a rat alveolar cell line (L2) in response to overnight exposure to either E2 or P4. In contrast to P4, E2 increased ENaC channel activity ( NPo) through an increase in channel open probability ( Po) and an increased number of patches with observable channel activity. Apical plasma membrane abundance of the ENaC α-subunit (αENaC) more than doubled in response to E2 as determined by cell surface biotinylation. αENaC membrane abundance was approximately threefold greater in lungs from female rats in proestrus, when serum E2 is greatest, compared with diestrus, when it is lowest. Our results also revealed a significant role for the G protein-coupled estrogen receptor (Gper) to mediate E2's effects on ENaC. Overall, our results demonstrate that E2 signaling through Gper selectively activates alveolar ENaC through an effect on channel gating and channel density, the latter via greater trafficking of channels to the plasma membrane. The results presented herein implicate E2-mediated regulation of alveolar sodium channels in the sex differences observed in the pathogenesis of several pulmonary diseases.

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Ryanodine receptor dysfunction and triggered activity in the heart

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00924.2006 ◽

2007 ◽

Vol 292 (5) ◽

pp. H2144-H2151 ◽

Cited By ~ 21

Author(s):

Rodolphe P. Katra ◽

Toshiyuki Oya ◽

Gregory S. Hoeker ◽

Kenneth R. Laurita

Keyword(s):

Ryanodine Receptor ◽

Transmembrane Potential ◽

Calcium Release ◽

Left Ventricular ◽

Calcium Handling ◽

Adrenergic Stimulation ◽

Triggered Activity ◽

Multiple Regions ◽

Rapid Pacing ◽

Almost All

Arrhythmogenesis has been increasingly linked to cardiac ryanodine receptor (RyR) dysfunction. However, the mechanistic relationship between abnormal RyR function and arrhythmogenesis in the heart is not clear. We hypothesize that, under abnormal RyR conditions, triggered activity will be caused by spontaneous calcium release (SCR) events that depend on transmural heterogeneities of calcium handling. We performed high-resolution optical mapping of intracellular calcium and transmembrane potential in the canine left ventricular wedge preparation ( n = 28). Rapid pacing was used to initiate triggered activity under normal and abnormal RyR conditions induced by FKBP12.6 dissociation and β-adrenergic stimulation (20–150 μM rapamycin, 0.2 μM isoproterenol). Under abnormal RyR conditions, almost all preparations experienced SCRs and triggered activity, in contrast to control, rapamycin, or isoproterenol conditions alone. Furthermore, under abnormal RyR conditions, complex arrhythmias (monomorphic and polymorphic tachycardia) were commonly observed. After washout of rapamycin and isoproterenol, no triggered activity was observed. Surprisingly, triggered activity and SCRs occurred preferentially near the epicardium but not the endocardium ( P < 0.01). Interestingly, the occurrence of triggered activity and SCR events could not be explained by cytoplasmic calcium levels, but rather by fast calcium reuptake kinetics. These data suggest that, under abnormal RyR conditions, triggered activity is caused by multiple SCR events that depend on the faster calcium reuptake kinetics near the epicardium. Furthermore, multiple regions of SCR may be a mechanism for multifocal arrhythmias associated with RyR dysfunction.

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Axons provide the secretory machinery for trafficking of voltage-gated sodium channels in peripheral nerve

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1514943113 ◽

2016 ◽

Vol 113 (7) ◽

pp. 1823-1828 ◽

Cited By ~ 23

Author(s):

Carolina González ◽

José Cánovas ◽

Javiera Fresno ◽

Eduardo Couve ◽

Felipe A. Court ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Sodium Channel ◽

Sodium Channels ◽

Neural Function ◽

Axonal Membrane ◽

Local Synthesis ◽

Voltage Gated ◽

Voltage Gated Sodium Channels

The regulation of the axonal proteome is key to generate and maintain neural function. Fast and slow axoplasmic waves have been known for decades, but alternative mechanisms to control the abundance of axonal proteins based on local synthesis have also been identified. The presence of the endoplasmic reticulum has been documented in peripheral axons, but it is still unknown whether this localized organelle participates in the delivery of axonal membrane proteins. Voltage-gated sodium channels are responsible for action potentials and are mostly concentrated in the axon initial segment and nodes of Ranvier. Despite their fundamental role, little is known about the intracellular trafficking mechanisms that govern their availability in mature axons. Here we describe the secretory machinery in axons and its contribution to plasma membrane delivery of sodium channels. The distribution of axonal secretory components was evaluated in axons of the sciatic nerve and in spinal nerve axons after in vivo electroporation. Intracellular protein trafficking was pharmacologically blocked in vivo and in vitro. Axonal voltage-gated sodium channel mRNA and local trafficking were examined by RT-PCR and a retention-release methodology. We demonstrate that mature axons contain components of the endoplasmic reticulum and other biosynthetic organelles. Axonal organelles and sodium channel localization are sensitive to local blockade of the endoplasmic reticulum to Golgi transport. More importantly, secretory organelles are capable of delivering sodium channels to the plasma membrane in isolated axons, demonstrating an intrinsic capacity of the axonal biosynthetic route in regulating the axonal proteome in mammalian axons.

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TMEM110 regulates the maintenance and remodeling of mammalian ER–plasma membrane junctions competent for STIM–ORAI signaling

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1521924112 ◽

2015 ◽

Vol 112 (51) ◽

pp. E7083-E7092 ◽

Cited By ~ 39

Author(s):

Ariel Quintana ◽

Vangipurapu Rajanikanth ◽

Suzette Farber-Katz ◽

Aparna Gudlur ◽

Chen Zhang ◽

...

Keyword(s):

Plasma Membrane ◽

Calcium Signaling ◽

Mammalian Cells ◽

Calcium Release ◽

Close Contact ◽

Transmembrane Protein ◽

Regulatory Protein ◽

Major Mechanism ◽

Genome Wide ◽

Core Elements

The stromal interaction molecule (STIM)–ORAI calcium release-activated calcium modulator (ORAI) pathway controls store-dependent calcium entry, a major mechanism of physiological calcium signaling in mammalian cells. The core elements of the pathway are the regulatory protein STIM1, located in the endoplasmic reticulum (ER) membrane, the calcium channel ORAI1 in the plasma membrane, and sites of close contact between the ER and the plasma membrane that permit the two proteins to interact. Research on calcium signaling has centered on STIM1, ORAI1, and a few proteins that directly modulate STIM–ORAI function. However, little is known about proteins that organize ER–plasma membrane junctions for STIM–ORAI-dependent calcium signaling. Here, we report that an ER-resident membrane protein identified in a previous genome-wide RNAi screen, transmembrane protein 110 (TMEM110), regulates the long-term maintenance of ER–plasma membrane junctions and the short-term physiological remodeling of the junctions during store-dependent calcium signaling.

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Membrane junctions in xenopus eggs: their distribution suggests a role in calcium regulation

The Journal of Cell Biology ◽

10.1083/jcb.96.4.1159 ◽

1983 ◽

Vol 96 (4) ◽

pp. 1159-1163 ◽

Cited By ~ 92

Author(s):

DM Gardiner ◽

RD Grey

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Intracellular Calcium ◽

Calcium Release ◽

Egg Activation ◽

Electron Dense Material ◽

Intracellular Calcium Release ◽

Cortical Endoplasmic Reticulum ◽

Center Distance ◽

Lines Of Evidence

We have observed the presence of membrane junctions formed between the plasma membrane and cortical endoplasmic reticulum of mature, unactivated eggs of xenopus laevis. The parallel, paired membranes of the junction are separated by a 10-mn gap within which electron-dense material is present. This material occurs in patches with an average center-to-center distance of approximately 30 nm. These junctions are rare in immature (but fully grown) oocytes (approximately 2 percent of the plasma membrane is associated with junctions) and increase dramatically during progesterone-induced maturation. Junctions in the mature, unactivated egg are two to three times more abundant in the animal hemisphere (25-30 percent of the plasma membrane associated with junction) as compared with the vegetal hemisphere (10-15 percent). Junction density decreases rapidly to values characteristic of immature oocytes in response to egg activation. The plasma membrane-ER junctions of xenopus eggs are strikingly similar in structure to membrane junctions in muscle cells thought to be essential in the triggering of intracellular calcium release from the sarcoplasmic reticulum. In addition, the junctions' distinctive, animal-vegetal polarity of distribution, their dramatic appearance during maturation, and their disapperance during activation are correlated with previously documented patterns of calcium-mediated events in anuran eggs. We discuss several lines of evidence supporting the hypothesis that these junctions in xenopus eggs are sites that transduce extracellular events into intracellular calcium release during fertilization and activation of development.

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Optical investigations reveal the effects of 2-aminoethyldiphenyl borate on STIM1 puncta formation

Journal of Innovative Optical Health Sciences ◽

10.1142/s1793545818500037 ◽

2018 ◽

Vol 11 (02) ◽

pp. 1850003 ◽

Cited By ~ 2

Author(s):

Tao Yu ◽

Shangbin Chen ◽

Jingying Pan ◽

Conglin Su ◽

Jun He

Keyword(s):

Plasma Membrane ◽

Inhibitory Action ◽

Calcium Release ◽

Confocal Imaging ◽

Text Entry ◽

Inhibitory Mechanism ◽

Inhibitory Effects ◽

Inhibition Effect ◽

Imaging Results ◽

And Function

2-Aminoethyldiphenyl borate (2-APB) is the most commonly used pharmacological agent in the study of calcium release-activated channels (CRACs); however, its inhibitory mechanism to CRACs remains unclear. To address this issue, we systematically employed confocal imaging, dual-wavelength excitation photometry and FRET to examine the effects of 2-APB on the dynamic activities and function of STIM1 and Orai1, two key components of CRACs. Imaging results support that there are two signaling pathways (Orai1-independent and Orai1-dependent) for the formation of STIM1 puncta. 2-APB could dose dependently block Orai1-independent but not Orai1-dependent STIM1 puncta formation, despite its obvious inhibition effect on store-operated Ca[Formula: see text] entry (SOCE). In addition, we found that although 2-APB could not visibly alter near plasma membrane CAD-eYFP localization, it could completely block CAD-YFP-induced constitutive Ca[Formula: see text] entry and promote the interaction between Orai1 and CAD by FRET measurements. Therefore, we proposed that inhibitory action of 2-APB on SOCE might attribute to its direct inhibitory effects on Orai1 channel itself, but not the interference on puncta formation between STIM1 and Orai1.

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Evidence for the involvement of microtubules, ER, and kinesin in the cortical rotation of fertilized frog eggs.

The Journal of Cell Biology ◽

10.1083/jcb.114.5.1017 ◽

1991 ◽

Vol 114 (5) ◽

pp. 1017-1028 ◽

Cited By ~ 65

Author(s):

E Houliston ◽

R P Elinson

Keyword(s):

Cell Cycle ◽

Plasma Membrane ◽

Sea Urchin ◽

Rana Pipiens ◽

Cortical Microtubules ◽

Cell Biol ◽

Vegetal Cortex ◽

Almost All ◽

Cytoplasmic Side ◽

Cytokeratin Filaments

During the first cell cycle, the vegetal cortex of the fertilized frog egg is translocated over the cytoplasm. This process of cortical rotation creates regional cytoplasmic differences important in later development, and appears to involve an array of aligned microtubules that forms transiently beneath the vegetal cortex. We have investigated how these microtubules might be involved in generating movement by analyzing isolated cortices and sections of Xenopus laevis and Rana pipiens eggs. First, the polarity of the cortical microtubules was determined using the "hook" assay. Almost all microtubules had their plus ends pointing in the direction of cortical rotation. Secondly, the association of microtubules with other cytoplasmic elements was examined. Immunofluorescence revealed that cytokeratin filaments coalign with the microtubules. The timing of their appearance and their position on the cytoplasmic side of the microtubules suggested that they are not involved directly in generating movement. ER was visualized with the dye DiIC16(3) and by immunofluorescence with anti-BiP (Bole, D. G., L. M. Hendershot, and J. F. Kearney, 1986. J. Cell Biol. 102:1558-1566). One layer of ER was found closely underlying the plasma membrane at all times. An additional, deeper layer formed in association with the microtubules of the array. Antibodies to sea urchin kinesin (Ingold, A. L., S. A. Cohn, and J. M. Scholey. 1988. J. Cell Biol. 107:2657-2667) detected antigens associated with both the ER and microtubules. On immunoblots they recognized microtubule associated polypeptide(s) of approximately 115 kD from Xenopus eggs. These observations are consistent with a role for kinesin in creating movement between the microtubules and ER, which leads in turn to the cortical rotation.

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