scholarly journals [Ca2+]i oscillations in human sperm are triggered in the flagellum by membrane potential-sensitive activity of CatSper

2020 ◽  
Author(s):  
Elis Torrezan-Nitao ◽  
Sean G Brown ◽  
Esperanza Mata-Martínez ◽  
Claudia L Treviño ◽  
Christopher Barratt ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Large Scale ◽  
Conflicts Of Interest ◽  
Human Sperm ◽  
Equilibrium Potential ◽  
Current Clamp ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current

Abstract STUDY QUESTION How are progesterone (P4)-induced repetitive intracellular Ca2+ concentration ([Ca2+]i) signals (oscillations) in human sperm generated? SUMMARY ANSWER P4-induced [Ca2+]i oscillations are generated in the flagellum by membrane potential (Vm)-sensitive Ca2+-influx through CatSper channels. WHAT IS KNOWN ALREADY A subset of human sperm display [Ca2+]i oscillations that regulate flagellar beating and acrosome reaction. Although pharmacological manipulations indicate involvement of stored Ca2+ in these oscillations, influx of extracellular Ca2+ is also required. STUDY DESIGN, SIZE, DURATION This was a laboratory study that used >20 sperm donors and involved more than 100 separate experiments and analysis of more than 1000 individual cells over a period of 2 years. PARTICIPANTS/MATERIALS, SETTING, METHODS Semen donors and patients were recruited in accordance with local ethics approval from Birmingham University and Tayside ethics committees. [Ca2+]i responses and Vm of individual cells were examined by fluorescence imaging and whole-cell current clamp. MAIN RESULTS AND THE ROLE OF CHANCE P4-induced [Ca2+]i oscillations originated in the flagellum, spreading to the neck and head (latency of 1–2 s). K+-ionophore valinomycin (1 µM) was used to investigate the role of membrane potential (Vm). Direct assessment by whole-cell current-clamp confirmed that Vm in valinomycin-exposed cells was determined primarily by K+ equilibrium potential (EK) and was rapidly ‘reset’ upon manipulation of [K+]o. Pre-treatment of sperm with valinomycin ([K+]o = 5.4 mM) had no effect on the P4-induced [Ca2+] transient (P = 0.95; eight experiments), but application of valinomycin to P4-pretreated sperm suppressed activity in 82% of oscillating cells (n = 257; P = 5 × 10−55 compared to control) and significantly reduced both the amplitude and frequency of persisting oscillations (P = 0.0001). Upon valinomycin washout, oscillations re-started in most cells. When valinomycin was applied in saline with elevated [K+], the inhibitory effect of valinomycin was reduced and was dependent on EK (P = 10−25). Amplitude and frequency of [Ca2+]i oscillations that persisted in the presence of valinomycin showed similar sensitivity to EK (P < 0.01). The CatSper inhibitor RU1968 (4.8 and 11 µM) caused immediate and reversible arrest of activity in 36% and 96% of oscillating cells, respectively (P < 10−10). Quinidine (300 µM) which blocks the sperm K+ current (IKsper) completely, inhibited [Ca2+]i oscillations. LARGE SCALE DATA N/A LIMITATIONS, REASONS FOR CAUTION This was an in-vitro study and caution must be taken when extrapolating these results to in-vivo regulation of sperm. WIDER IMPLICATIONS OF THE FINDINGS [Ca2+]i oscillations in human sperm are functionally important and their absence is associated with failed fertilisation at IVF. The data reported here provide new understanding of the mechanisms that underlie the regulation and generation (or failure) of these oscillations. STUDY FUNDING/COMPETING INTEREST(S) E.T.-N. was in receipt of a postgraduate scholarship from the CAPES Foundation (Ministry of Education, Brazil). E.M-M received travel funds from the Programa de Apoyo a los Estudios de Posgrado (Maestria y Doctorado en Ciencias Bioquimicas-Universidad Autonoma de Mexico). SGB and CLRB are recipients of a Chief Scientist Office (NHS Scotland) grant TCS/17/28. The authors have no conflicts of interest.

Metabolic communication from cardiac myocytes to vascular endothelial cells

2005 ◽  
Vol 288 (5) ◽  
pp. H2232-H2237 ◽  
Author(s):  
Anna K. Brzezinska ◽  
Daphne Merkus ◽  
William M. Chilian
Keyword(s):  
Endothelial Cells ◽  
Membrane Potential ◽  
Cardiac Myocytes ◽  
Vascular Tone ◽  
Vascular Endothelial Cells ◽  
Resting Membrane Potential ◽  
Vascular Endothelial ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current

The endothelium releases substances that affect both vascular and cardiac myocytes. However, under conditions of augmented metabolic demands and cardiac work, signals from the cardiac myocytes may be critical for the endothelium to fulfill its secretory and regulatory function in the vascular bed. Therefore, we hypothesized that cardiac myocytes produce substances that alter the resting membrane potential of endothelial cells and thus vascular tone. Isolated rat cardiac myocytes were electrically stimulated at the rate of 0 and 400 beats/min (Po2 = 150 mmHg), and supernatants were collected from each group (Sup-0; control) and (Sup-400) and used within 6 mo. These supernatants were applied to human coronary endothelial cells that were subsequently analyzed by using the whole cell and cell-attached patch-clamp modes. Sup-0 had no effect on the whole cell current and the zero-current potential. The Sup-0 from myocytes treated with aprotinin, an inhibitor of kallikrein and serine protease, reduced whole cell current between −120 and −60 mV. Sup-400 depolarized endothelial cells from the resting membrane potential of −45 to −5 mV ( P < 0.05), increased the magnitude of an inward current, and activated an outward current. Moreover, Sup-400 cells assayed in cell-attached patches increased single channel amplitude and the probability of a channel being in the open state. These effects were reversed by the Sup-400 from aprotinin-treated cells. We conclude that under certain metabolic conditions, isolated cardiac myocytes produce and release vasoactive substances that alter the function of K+ channels in vascular endothelial cells. Thus cardiac myocytes seem to communicate metabolic information to the endothelium, which could potentially influence vascular tone.

Contribution of the KCa channel to membrane potential and O2 sensitivity is decreased in an ovine PPHN model

2002 ◽  
Vol 283 (5) ◽  
pp. L1103-L1109 ◽  
Author(s):  
Andrea Olschewski ◽  
Zhigang Hong ◽  
Bradley C. Linden ◽  
Valerie A. Porter ◽  
E. Kenneth Weir ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Membrane Potential ◽  
Ductus Arteriosus ◽  
Ovine Model ◽  
Whole Cell ◽  
Pulmonary Vasodilation ◽  
Cell Current ◽  
Pulmonary Artery Smooth Muscle ◽  
Perforated Patch Clamp ◽  
Whole Cell Current

Ca2+-sensitive K+(KCa) channels play an important role in mediating perinatal pulmonary vasodilation. We hypothesized that lung KCa channel function may be decreased in persistent pulmonary hypertension of the newborn (PPHN). To test this hypothesis, pulmonary artery smooth muscle cells (PASMC) were isolated from fetal lambs with severe pulmonary hypertension induced by ligation of the ductus arteriosus in fetal lambs at 125–128 days gestation. Fetal lambs were killed after pulmonary hypertension had been maintained for at least 7 days. Age-matched, sham-operated animals were used as controls. PASMC K+ currents and membrane potentials were recorded using amphotericin B-perforated patch-clamp techniques. The increase in whole cell current normally seen in response to normoxia was decreased (333.9 ± 63.6% in control vs. 133.1 ± 16.0% in hypertensive fetuses). The contribution of the KCachannel to the whole cell current was diminished in hypertensive, compared with control, fetal PASMC. In PASMC from hypertensive fetuses, a change from hypoxia to normoxia caused no change in membrane potential compared with a −14.6 ± 2.8 mV decrease in membrane potential in PASMC from control animals. In PASMC from animals with pulmonary hypertension, 4-aminopyridine (4-AP) caused a larger depolarization than iberiotoxin, whereas in PASMC from control animals, iberiotoxin caused a larger depolarization than 4-AP. These data confirm the hypothesis that the contribution of the KCachannel to membrane potential and O2 sensitivity is decreased in an ovine model of PPHN, and this may contribute to the abnormal perinatal pulmonary vasoreactivity associated with PPHN.

Activation of whole cell currents in isolated human jejunal circular smooth muscle cells by carbon monoxide

1993 ◽  
Vol 264 (6) ◽  
pp. G1184-G1189 ◽  
Author(s):  
G. Farrugia ◽  
W. A. Irons ◽  
J. L. Rae ◽  
M. G. Sarr ◽  
J. H. Szurszewski
Keyword(s):  
Carbon Monoxide ◽  
Smooth Muscle ◽  
Membrane Potential ◽  
Muscle Cells ◽  
Patch Clamp Technique ◽  
Current Increase ◽  
Membrane Hyperpolarization ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current

Carbon monoxide (CO) is a low molecular weight oxide produced endogenously from fatty acids and heme protein. A physiological role for CO has been suggested for vascular smooth muscle, hemostasis, and olfactory neurons, but direct evidence is lacking. Heme oxygenase, which catalyzes the formation of CO from heme proteins, is present in small intestinal smooth muscle. The effect of 1% CO on whole cell currents in normal human jejunal circular muscle cells was studied with the use of a perforated patch-clamp technique. A 1% CO-containing Krebs solution caused an initial and transient increase in whole cell current in 20 of 22 cells tested (175 +/- 40%, mean +/- SE) and a transient hyperpolarization (15.6 +/- 3.6 mV, mean +/- SE) of the membrane potential. During prolonged recordings, 1% CO evoked ongoing cyclic increases and decreases in the whole cell current. Each current increase was accompanied by a sharp membrane hyperpolarization. These data suggest that CO may modulate whole cell potassium current and membrane potential.

scholarly journals Modulation of inhibitory and excitatory fast neurotransmission in the rat CNS by heavy water (D2O)

2015 ◽  
Vol 114 (2) ◽  
pp. 1109-1118 ◽  
Author(s):  
Masahito Wakita ◽  
Naoki Kotani ◽  
Kiyomitsu Shoudai ◽  
Toshitaka Yamaga ◽  
Norio Akaike
Keyword(s):  
Heavy Water ◽  
Deuterium Oxide ◽  
Dependent Manner ◽  
General Anesthetics ◽  
Concentration Dependent Manner ◽  
Excitatory Postsynaptic Currents ◽  
Whole Cell ◽  
Postsynaptic Currents ◽  
Cell Current ◽  
Whole Cell Current

The effects of heavy water (deuterium oxide, D2O) on GABAergic and glutamatergic spontaneous and evoked synaptic transmission were investigated in acute brain slice and isolated “synaptic bouton” preparations of rat hippocampal CA3 neurons. The substitution of D2O for H2O reduced the frequency and amplitude of GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) in a concentration-dependent manner but had no effect on glutamatergic spontaneous excitatory postsynaptic currents (sEPSCs). In contrast, for evoked synaptic responses in isolated neurons, the amplitude of both inhibitory and excitatory postsynaptic currents (eIPSCs and eEPSCs) was decreased in a concentration-dependent manner. This was associated with increases of synaptic failure rate ( Rf) and paired-pulse ratio (PPR). The effect was larger for eIPSCs compared with eEPSCs. These results clearly indicate that D2O acts differently on inhibitory and excitatory neurotransmitter release machinery. Furthermore, D2O significantly suppressed GABAA receptor-mediated whole cell current ( IGABA) but did not affect glutamate receptor-mediated whole cell current ( IGlu). The combined effects of D2O at both the pre- and postsynaptic sites may explain the greater inhibition of eIPSCs compared with eEPSCs. Finally, D2O did not enhance or otherwise affect the actions of the general anesthetics nitrous oxide and propofol on spontaneous or evoked GABAergic and glutamatergic neurotransmissions, or on IGABA and IGlu. Our results suggest that previously reported effects of D2O to mimic and/or modulate anesthesia potency result from mechanisms other than modulation of GABAergic and glutamatergic neurotransmission.

Potassium currents in mammalian and avian isolated type I semicircular canal hair cells

1994 ◽  
Vol 71 (1) ◽  
pp. 317-329 ◽  
Author(s):  
K. J. Rennie ◽  
M. J. Correia
Keyword(s):  
Membrane Potential ◽  
Hair Cells ◽  
Potassium Current ◽  
Columba Livia ◽  
Membrane Properties ◽  
Delayed Rectifier ◽  
Type I ◽  
Type Ii ◽  
Current Clamp ◽  
Whole Cell

1. Type I vestibular hair cells were isolated from the cristae ampullares of the semicircular canals of the Mongolian gerbil (Meriones unguiculatus) and the white king pigeon (Columba livia). Dissociated type I cells were distinguished from type II hair cells by their neck to plate ratio (NPR) and their characteristic amphora shape. 2. The membrane properties of gerbil and pigeon type I hair cells were studied in whole-cell voltage- and current-clamp using the perforated patch technique with amphotericin B as the perforating agent. 3. In whole-cell current-clamp, the average zero-current potential, Vz, measured for pigeon type I hair cells, was -70 +/- 7 (SD) mV (n = 18) and -71 +/- 11 mV (n = 83) for gerbil type I hair cells. 4. At Vz, for both gerbil and pigeon type I hair cells, a potassium current (IKI) was > or = 50% activated. This current deactivated rapidly when the membrane potential was hyperpolarized below -90 mV. 5. IKI was blocked by externally applied 4-aminopyridine (4-AP) (5 mM) and by internally applied 20 mM tetraethylammonium (TEA). It was also reduced when 4 mM barium was present in the external solution. The degree of block by barium increased as the membrane potential became more positive. External cesium (5 mM) blocked the inward component of IKI. When IKI was pharmacologically blocked, Vz depolarized by approximately 40 mV. Therefore IKI appears to be a delayed rectifier and to set the more negative Vz noted for isolated type I hair cells when compared to isolated type II hair cells, which do not have IKI. 6. A second, smaller potassium current was present at membrane potential depolarizations above -40 mV. This current was blocked by 30-50 mM, externally applied TEA, 100 microM quinidine, 100 nM apamin, but not 100 nM charybdotoxin, indicating that this is a calcium-activated potassium current, IK(Ca), different from the maxi-K calcium-activated potassium current found in most other hair cells.

Monozygotic Twins Display Different Minor Histocompatibility Antigens.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4324-4324
Author(s):  
Miroslaw Markiewicz ◽  
Urszula Siekiera ◽  
Monika Dzierzak Mietla ◽  
Agnieszka Karolczyk ◽  
Tomasz Kruzel ◽  
...  
Keyword(s):  
Large Scale ◽  
Hematological Malignancies ◽  
Conflicts Of Interest ◽  
Monozygotic Twins ◽  
Genotypic Diversity ◽  
Histocompatibility Antigens ◽  
Phenotypic Differences ◽  
Minor Histocompatibility Antigens ◽  
Genes Encoding

Abstract Abstract 4324 Introduction Albeit it is generally presumed that monozygotic twins are genetically identical and that phenotypic differences between twins are mainly due to environmental factors, large-scale variation in copy number of DNA segments recently evidenced by Bruder et al. (AJHG, 2008) showed presence of genotypic diversity in monozygotic twins. The rationale of this study was to test whether monozygotic twins display disparities of minor Histocompatibility antigens (mHags) which may play role in syngenic HCT. We and others have previously shown that mHags constitute an important immunogenetic factor influencing immune responses following transplantation from HLA-matched allogeneic donors. Patients and Methods mHags HA-1, HA-2, HA-3, HA-8, HB-1, ACC-1, ACC-2, HwA-9, HwA-10, UGT2B17, HY genotypes were defined with use of Dynal AllSet kits by PCR-SSP method in secured DNA samples from 3 monozygotic twins pairs aged 34, 24 and 28, who underwent syngenic allo-HCTs due to different hematological malignancies (NHL, CML, AML) in the Department of Hematology and BMT in Katowice, Poland in years 2000-2004. Results In 2 out of 3 syngenic pairs we have found differences in genes encoding mHags: different allele of EB-1 was present in one pair (NHL) (recipient HH, donor HY), and two different alleles of HwA-9 (RR, RG) and HwA-10 (**, R*) were present in second pair (CML). No differences in mHags were observed in the third pair (AML). Conclusions Our results question the long-standing belief that monozygotic twins are genetically identical and open up a possibility to further study the role of disparate mHags in disease and transplantation. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Ionflux: A Microfluidic Approach to Ensemble Recording and Block of Whole-Cell Current from Voltage-Gated Ion Channels

2010 ◽  
Vol 98 (3) ◽  
pp. 194a ◽  
Author(s):  
C. Ian Spencer ◽  
Nianzhen Li ◽  
Juliette Johnson ◽  
Qin Chen ◽  
Cristian Ionescu-Zanetti
Keyword(s):  
Ion Channels ◽  
Whole Cell ◽  
Voltage Gated ◽  
Cell Current ◽  
Voltage Gated Ion Channels ◽  
Whole Cell Current ◽  
Gated Ion Channels

scholarly journals The Effectiveness in Activating M-Type K+ Current Produced by Solifenacin ([(3R)-1-azabicyclo[2.2.2]octan-3-yl] (1S)-1-phenyl-3,4-dihydro-1H-isoquinoline-2-carboxylate): Independent of Its Antimuscarinic Action

2021 ◽  
Vol 22 (22) ◽  
pp. 12399
Author(s):  
Hsin-Yen Cho ◽  
Tzu-Hsien Chuang ◽  
Sheng-Nan Wu
Keyword(s):  
Single Channel ◽  
Ionic Currents ◽  
Reaction Scheme ◽  
Gh3 Cells ◽  
Whole Cell ◽  
Type K ◽  
Cell Current ◽  
K Current ◽  
Subsequent Addition ◽  
Whole Cell Current

Solifenacin (Vesicare®, SOL), known to be a member of isoquinolines, is a muscarinic antagonist that has anticholinergic effect, and it has been beneficial in treating urinary incontinence and neurogenic detrusor overactivity. However, the information regarding the effects of SOL on membrane ionic currents is largely uncertain, despite its clinically wide use in patients with those disorders. In this study, the whole-cell current recordings revealed that upon membrane depolarization in pituitary GH3 cells, the exposure to SOL concentration-dependently increased the amplitude of M-type K+ current (IK(M)) with effective EC50 value of 0.34 μM. The activation time constant of IK(M) was concurrently shortened in the SOL presence, hence yielding the KD value of 0.55 μM based on minimal reaction scheme. As cells were exposed to SOL, the steady-state activation curve of IK(M) was shifted along the voltage axis to the left with no change in the gating charge of the current. Upon an isosceles-triangular ramp pulse, the hysteretic area of IK(M) was increased by adding SOL. As cells were continually exposed to SOL, further application of acetylcholine (1 μM) failed to modify SOL-stimulated IK(M); however, subsequent addition of thyrotropin releasing hormone (TRH, 1 μM) was able to counteract SOL-induced increase in IK(M) amplitude. In cell-attached single-channel current recordings, bath addition of SOL led to an increase in the activity of M-type K+ (KM) channels with no change in the single channel conductance; the mean open time of the channel became lengthened. In whole-cell current-clamp recordings, the SOL application reduced the firing of action potentials (APs) in GH3 cells; however, either subsequent addition of TRH or linopirdine was able to reverse SOL-mediated decrease in AP firing. In hippocampal mHippoE-14 neurons, the IK(M) was also stimulated by adding SOL. Altogether, findings from this study disclosed for the first time the effectiveness of SOL in interacting with KM channels and hence in stimulating IK(M) in electrically excitable cells, and this noticeable action appears to be independent of its antagonistic activity on the canonical binding to muscarinic receptors expressed in GH3 or mHippoE-14 cells.

scholarly journals On the Contribution of the First Transmembrane Domain to Whole-Cell Current through an ATP-Gated Ionotropic P2X Receptor

2001 ◽  
Vol 21 (16) ◽  
pp. 5885-5892 ◽  
Author(s):  
William R. Haines ◽  
Mark M. Voigt ◽  
Keisuke Migita ◽  
Gonzalo E. Torres ◽  
Terrance M. Egan
Keyword(s):  
Transmembrane Domain ◽  
P2x Receptor ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current
Sign in / Sign up

Export Citation Format

Share Document