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Dierk Thomas - One of the best experts on this subject based on the ideXlab platform.
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altered hcn4 channel c linker interaction is associated with familial tachycardia bradycardia syndrome and atrial fibrillation
European Heart Journal, 2013Co-Authors: Nana Duhme, Dierk Thomas, Patrick Schweizer, Rudiger Becker, Julian Schroter, Thomas R M Barends, Ilme Schlichting, Andreas Draguhn, Claus BruehlAbstract:Aims HCN4 channels are involved in generation, regulation, and stabilization of heart rhythm and channel dysfunction is associated with inherited sinus bradycardia. We asked whether dysfunctional HCN4 channels also contribute to the generation of cardiac tachyarrhythmias. Methods and results In a candidate gene approach, we screened 422 patients with atrial and/or ventricular tachyarrhythmias and detected a novel HCN4 gene mutation that replaced the positively charged lysine 530 with an asparagine (HCN4-K530N) in a highly conserved region of the C-linker. The index patient developed tachycardia–bradycardia syndrome and persistent atrial fibrillation (AF) in an age-dependent fashion. Pedigree analysis identified eight affected family members with a similar course of disease. Whole-cell patch clamp electrophysiology of HEK293 cells showed that homomeric mutant channels almost are indistinguishable from wild-type channels. In contrast, heteromeric channels composed of mutant and wild-type subunits displayed a significant hyperpolarizing shift in the half-maximal Activation Voltage. This may be caused by a shift in the equilibrium between the tonically inhibited nucleotide-free state of the C-terminal domain of HCN4 believed to consist of a ‘dimer of dimers’ and the activated ligand-bound tetrameric form, leading to an increased inhibition of activity in heteromeric channels. Conclusion Altered C-linker oligomerization in heteromeric channels is considered to promote familial tachycardia–bradycardia syndrome and persistent AF, indicating that f-channel dysfunction contributes to the development of atrial tachyarrhythmias.
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altered hcn4 channel c linker interaction is associated with familial tachycardia bradycardia syndrome and atrial fibrillation
Biophysical Journal, 2012Co-Authors: Patrick A Schweizer, Hugo A. Katus, Nana Duhme, Rudiger Becker, Ilme Schlichting, Andreas Draguhn, Claus Bruehl, Michael Koenen, Dierk ThomasAbstract:HCN channels underlie the pacemaker current If, involved in generation, regulation, and stabilization of sinus rhythm. HCN4 represents the dominant isotype in the sinoatrial node and channel dysfunction is associated with inherited sinus node bradycardia. Here, we report a previously undescribed HCN4 gene mutation that replaced the positively charged lysine 530 with an asparagine (HCN4-K530N) in a highly conserved region of the C-linker. Six members of a German family carrying the HCN4-K530N mutation developed tachycardia-bradycardia syndrome and persistent atrial fibrillation in an age-dependent fashion. HEK293 cell recordings using whole-cell patch clamp electrophysiology revealed that homomeric HCN4-K530N mutant and wild type channels activate at similar potentials and respond equally upon binding of cAMP. Heteromeric channels, in contrast, showed a significant hyperpolarizing shift in the half-maximal Activation Voltage. Moreover, the effect of cAMP on channel Activation and deActivation properties was significantly increased in heteromeric channels. A comparison of mutant and wild type C-linker domain models suggests that altered subunit interactions between the A’ and B’ helices and the C’ and D’ helices of the neighboring subunit may change the intersubunit structural dynamics in heterotetramers of HCN4 wild type and mutant subunits, enforcing inhibition of channel activity by the nucleotide free cyclic nucleotide binding domain in the heterozygous situation. Thus, altered interaction of side chains is tolerated in homomeric mutant channels but interferes with wild type subunits in the heteromeric complex leading to f-channel dysfunction that promotes the development of tachycardia-bradycardia syndrome and persistent atrial fibrillation.
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Anticholinergic antiparkinson drug orphenadrine inhibits HERG channels: Block attenuation by mutations of the pore residues Y652 or F656
Naunyn-Schmiedeberg's Archives of Pharmacology, 2007Co-Authors: Eberhard P Scholz, Franziska M. Konrad, Daniel L. Weiss, Claudia Kiesecker, Martin Kulzer, S. Kathöfer, Edgar Zitron, Dierk Thomas, Ramona Bloehs, Alexander BauerAbstract:Abstract The anticholinergic antiparkinson drug orphenadrine is an antagonist at central and peripheral muscarinic receptors. Orphenadrine intake has recently been linked to QT prolongation and Torsade-de-Pointes tachycardia. So far, inhibitory effects on I Kr or cloned HERG channels have not been examined. HERG channels were heterologously expressed in a HEK 293 cell line and in Xenopus oocytes and HERG current was measured using the whole cell patch clamp and the double electrode Voltage clamp technique. Orphenadrine inhibits cloned HERG channels in a concentration dependent manner, yielding an IC50 of 0.85 ++M in HEK cells. Onset of block is fast and reversible upon washout. Orphenadrine does not alter the half-maximal Activation Voltage of HERG channels. There is no shift of the half-maximal steady-state-inActivation Voltage. Time constants of direct channel inActivation are not altered significantly and there is no use-dependence of block. HERG blockade is attenuated significantly in mutant channels lacking either of the aromatic pore residues Y652 and F656. In conclusion, we show that the anticholinergic agent orphenadrine is an antagonist at HERG channels. These results provide a novel molecular basis for the reported proarrhythmic side effects of orphenadrine
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acute effects of dronedarone on both components of the cardiac delayed rectifier k current herg and kvlqt1 mink potassium channels
British Journal of Pharmacology, 2003Co-Authors: Dierk Thomas, S. Kathöfer, Edgar Zitron, Wei Zhang, Christoph A. Karle, Wolfgang Schoels, Hugo A. Katus, Volker A. W. Kreye, Anna-britt Wimmer, Johann KiehnAbstract:Dronedarone is a noniodinated benzofuran derivative that has been synthesized to overcome the limiting iodine-associated adverse effects of the potent antiarrhythmic drug amiodarone. In this study, the acute electrophysiological effects of dronedarone on repolarizing potassium channels were investigated to determine the class III antiarrhythmic action of this compound. HERG and KvLQT1/minK potassium channels conduct the delayed rectifier potassium current IK in human heart, being a primary target for class III antiarrhythmic therapy. HERG and KvLQT1/minK were expressed heterologously in Xenopus laevis oocytes, and the respective potassium currents were recorded using the two-microelectrode Voltage-clamp technique. Dronedarone blocked HERG channels with an IC50 value of 9.2 microM and a maximum tail current reduction of 85.2%. HERG channels were blocked in the closed, open, and inactivated states. The half-maximal Activation Voltage was shifted by -6.1 mV, and HERG current block by dronedarone was Voltage-dependent, but not use-dependent. Dronedarone exhibited a weaker block of KvLQT1/minK currents (33.2% at 100 microM drug concentration), without causing significant changes in the corresponding current-Voltage relationships. In conclusion, these data demonstrate that dronedarone is an antagonist of cloned HERG potassium channels, with additional inhibitory effects on KvLQT1/minK currents at higher drug concentrations, providing a molecular mechanism for the class III antiarrhythmic action of the drug.
Won Sun Park - One of the best experts on this subject based on the ideXlab platform.
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the t type ca2 channel inhibitor mibefradil inhibits Voltage dependent k channels in rabbit coronary arterial smooth muscle cells
Journal of Pharmacological Sciences, 2012Co-Authors: Da Hye Hong, Youn Kyoung Son, Dongki Yang, Ilwhan Choi, Wonkyo Jung, Dae Joong Kim, Jin Han, Won Sun ParkAbstract:Abstract We examined the effects of mibefradil, a T-type Ca2+ channel inhibitor, on Voltage-dependent K+ (Kv) channels in rabbit coronary arterial smooth muscle cells using the whole-cell patch clamp technique. Mibefradil reduced the Kv current amplitude in a dose-dependent manner, with an apparent Kd value of 1.08 μM. Kv current inhibition by mibefradil was highly Voltage-dependent over the full Activation Voltage range (−30 to +10 mV). The decay rate of Kv channel inActivation was accelerated by mibefradil without altering the kinetics of current Activation. The rate constants of association and dissociation were 2.23 ± 0.07 μM−1·s−1 and 2.40 ± 0.42 s−1, respectively. Mibefradil had no significant effect on the steady-state Activation or inActivation curves. In the presence of mibefradil, the recovery time constant from inActivation was decreased, and the application of train pulses (1 or 2 Hz) increased mibefradil-induced Kv channel inhibition, suggesting that the inhibitory effects of mibefradil were use-dependent. The inhibitory effect of mibefradil on Kv channels was unaffected by extracellular Ca2+-free conditions. Moreover, the absence of ATP inside the pipette did not alter the blocking effect of mibefradil. Therefore, we suggest that mibefradil directly inhibited the Kv current, independently of Ca2+ channel inhibition.
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the protein kinase c inhibitor bisindolylmaleimide i inhibits Voltage dependent k channels in coronary arterial smooth muscle cells
Life Sciences, 2005Co-Authors: Won Sun Park, Youn Kyoung Son, Kyoung Sun Park, Hyang Ae Lee, Yung E EarmAbstract:Abstract We examined the effects of the protein kinase C (PKC) inhibitor, bisindolylmaleimide (BIM) (I), on Voltage-dependent K + (K V ) channels in rabbit coronary arterial smooth muscle cells using whole-cell patch clamp technique. BIM (I) reversibly and dose-dependently inhibited the K V currents with an apparent K d value of 0.27 μM. The inhibition of the K V current by BIM (I) was highly Voltage-dependent between −30 and +10 mV (Voltage range of channel Activation), and the additive inhibition of the K V current by BIM (I) was Voltage-dependence in the full Activation Voltage range. The rate constants of association and dissociation for BIM (I) were 18.4 μM −1 s −1 and 4.7 s −1 , respectively. BIM (I) had no effect on the steady-state Activation and inActivation of K V channels. BIM (I) caused use-dependent inhibition of K V current, which was consistent with the slow recovery from inActivation in the presence of BIM (I) (recovery time constants were 856.95 ± 282.6 ms for control, and 1806.38 ± 110.0 ms for 300 nM BIM (I)). ATP-sensitive K + (K ATP ), inward rectifier K + (K IR ), Ca 2+ -activated K + (BK Ca ) channels, which regulate the membrane potential and arterial tone, were not affected by BIM (I). The PKC inhibitor, chelerythrine, and protein kinase A (PKA) inhibitor, PKA-IP, had little effect on the K V current and did not significantly alter the inhibitory effects of BIM (I) on the K V current. These results suggest that BIM (I) inhibits K V channels in a phosphorylation-independent, and Voltage-, time- and use-dependent manner.
Michael Wissler - One of the best experts on this subject based on the ideXlab platform.
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characterization of dielectric elastomer actuators based on a visco hyperelastic film model
Smart Materials and Structures, 2007Co-Authors: Patrick Lochmatter, Gabor Kovacs, Michael WisslerAbstract:The electromechanical performance of planar dielectric elastomer (DE) actuators is predicted by applying a novel model for the mechanical behavior of visco-hyperelastic films such as VHB 4910 (manufactured by 3M). The electrostatic pressure was introduced in the film thickness direction to adapt the film model to DE actuators. Moreover, the actuator was embedded in an appropriate electrical supply circuit to account for the electrodynamic effects. The simulation of the active expansion of a biaxially prestrained, planar DE actuator configuration showed unstable deformation behavior under long-term Activation. For Activation Voltages exceeding a critical level, the active expansion thus became uncontrolled after some time. The model was also applied to a DE strip actuator configuration under sinusoidal electromechanical excitation. The influence of selected parameters on the overall actuator performance was thereby investigated. While the specific energy density increases with increasing amplitudes of the Activation Voltage and the stretch ratio, the optimum efficiency is predicted to lie at moderate electromechanical excitations.
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circular pre strained dielectric elastomer actuator modeling simulation and experimental verification
Smart Structures and Materials 2005: Electroactive Polymer Actuators and Devices (EAPAD), 2005Co-Authors: Michael Wissler, Edoardo Mazza, Gabor KovacsAbstract:Reliable models are required for the design and optimization of dielectric elastomer actuators. Thereby knowledge of the constitutive behavior of the elastomer is of crucial importance. In this work a pre-strained circular actuator made of a dielectric elastomer is investigated: constitutive models based on uniaxial data are verified by comparing calculation results with experimental observations. An analytical model is derived for the instantaneous response to an Activation Voltage in the pre-strained circular actuator and a finite element (FE) model is used to simulate the time dependent behavior. Hyperelastic models are used and three strain energy formulations (Yeoh, Ogden and Mooney-Rivlin) are compared in their predictive capabilities. The results of the calculations with the three strain energy forms differ significantly, although all forms were successfully fitted to the same uniaxial data set. Predictions of the actuator behavior based on the Yeoh form agree to a great extent with measurements of the response at different pre-strain levels and Activation Voltages.
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modeling of a pre strained circular actuator made of dielectric elastomers
Sensors and Actuators A-physical, 2005Co-Authors: Michael Wissler, Edoardo MazzaAbstract:Dielectric elastomers are used for the realization of actuators with large deformations and belong to the group of so-called electroactive polymers (EAP). Models are required for the design and optimization of EAP actuators. Thereby the constitutive behavior of the elastomer is of crucial importance and typically uniaxial experiments are performed in order to determine the mechanical properties of these materials. In this paper a pre-strained circular actuator made of a dielectric elastomer is investigated: constitutive models based on uniaxial data are verified by comparing calculation results with experimental observations. An analytical model is derived for the instantaneous response to an Activation Voltage in the pre-strained circular actuator and a finite element model is used to simulate the time-dependent behavior. Hyperelastic models are used and three strain energy formulations (Yeoh, Ogden and Mooney-Rivlin) are compared in their predictive capabilities. The results of the calculations with the three strain energy forms differ significantly, although all forms were successfully fitted to the same uniaxial data set. Predictions of the actuator behavior with the Yeoh form agree to a great extent with the measurements. The results of the present work show that the circular actuator set-up represents a valid model system for the characterization and optimization of the electromechanical behavior of dielectric elastomers.
Ian D Forsythe - One of the best experts on this subject based on the ideXlab platform.
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nitric oxide selectively suppresses ih currents mediated by hcn1 containing channels
The Journal of Physiology, 2015Co-Authors: Cornelia Koppscheinpflug, Beatrice M. Pigott, Ian D ForsytheAbstract:Key points The superior olivary complex (SOC) exhibits a spectrum of HCN1 and HCN2 subunit expression, which generate IH currents with fast and slow kinetics, respectively. Neuronal nitric oxide synthase (nNOS) was broadly distributed across the SOC. NO hyperpolarizes the half-Activation Voltage of HCN1-mediated currents and caused a slowing of the IH current kinetics in the respective nuclei (medial and lateral superior olives and superior paraolivary nucleus). This signalling was independent of cGMP. NO also caused a depolarizing shift in the half-Activation Voltage of HCN2-mediated IH currents, increasing Activation at resting potentials; this was cGMP-dependent. Thus, NO signalling suppressed fast HCN1-mediated currents and potentiated slow HCN2-mediated currents, modulating the overall kinetics and magnitude of the endogenous IH. Abstract Hyperpolarization-activated non-specific cation-permeable channels (HCN) mediate IH currents, which are modulated by cGMP and cAMP and by nitric oxide (NO) signalling. Channel properties depend upon subunit composition (HCN1–4 and accessory subunits) as demonstrated in expression systems, but physiological relevance requires investigation in native neurons with intact intracellular signalling. Here we use the superior olivary complex (SOC), which exhibits a distinctive pattern of HCN1 and HCN2 expression, to investigate NO modulation of the respective IH currents, and compare properties in wild-type and HCN1 knockout mice. The medial nucleus of the trapezoid body (MNTB) expresses HCN2 subunits exclusively, and sends inhibitory projections to the medial and lateral superior olives (MSO, LSO) and the superior paraolivary nucleus (SPN). In contrast to the MNTB, these target nuclei possess an IH with fast kinetics, and they express HCN1 subunits. NO is generated in the SOC following synaptic activity and here we show that NO selectively suppresses HCN1, while enhancing IH mediated by HCN2 subunits. NO hyperpolarizes the half-Activation of HCN1-mediated currents and slows the kinetics of native IH currents in the MSO, LSO and SPN. This modulation was independent of cGMP and absent in transgenic mice lacking HCN1. Independently, NO signalling depolarizes the half-Activation of HCN2-mediated IH currents in a cGMP-dependent manner. Thus, NO selectively suppresses fast HCN1-mediated IH and facilitates a slow HCN2-mediated IH, so generating a spectrum of modulation, dependent on the local expression of HCN1 and/or HCN2.
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nitric oxide signalling augments neuronal Voltage gated l type cav1 and p q type cav2 1 channels in the mouse medial nucleus of the trapezoid body
PLOS ONE, 2012Co-Authors: Adam J B Tozer, Ian D Forsythe, Joern R SteinertAbstract:Nitric Oxide (NO) is a diffusible second messenger that modulates ion channels, intrinsic excitability and mediates synaptic plasticity. In light of its activity-dependent generation in the principal neurons of the medial nucleus of the trapezoid body (MNTB), we have investigated its potential modulatory effects on native Voltage-gated calcium channels (CaV) within this nucleus. Whole-cell patch recordings were made from brain slices from P13–15 CBA mice. Slices were incubated with the inhibitor of neuronal nitric oxide synthase (nNOS) 7-nitroindazole (10 µM) and pharmacological blockers used to isolate Ca2+ current subtypes. Unpaired observations in the presence and absence of the NO-donors sodium nitroprusside (SNP, 100 µM) or Diethyl-ammonium-nonoate (DEA, 100 µM) were made to elucidate NO-dependent modulation of the expressed CaV subtypes. A differential effect of NO on the calcium channel subtypes was observed: CaV1 and CaV2.1 (L+R- and P/Q+R-type) conductances were potentiated, whereas N+R-type (CaV2.2) and R-type (CaV2.3) current amplitudes were unaffected. L+R-type currents increased from 0.36±0.04 nA to 0.64±0.11 nA and P/Q+R-type from 0.55±0.09 nA to 0.94±0.05 nA, thereby changing the balance and relative contribution of each subtype to the whole cell calcium current. In addition, N+R-type half-Activation Voltage was left shifted following NO exposure. NO-dependent modulation of P/Q+R and N+R-type, but not L+R-type, channels was removed by inhibition of soluble guanylyl cyclase (sGC) activity. This data demonstrates a differential effect of NO signalling on Voltage-gated calcium entry, by distinct NO-dependent pathways.
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facilitation of the presynaptic calcium current at an auditory synapse in rat brainstem
Web Science, 1998Co-Authors: Matthew F Cuttle, Tetsuhiro Tsujimoto, Ian D ForsytheAbstract:1. The presynaptic calcium current (IpCa) was recorded from the calyx of Held in rat brainstem slices using the whole-cell patch clamp technique. 2. Tetanic Activation of IpCa by 1 ms depolarizing Voltage steps markedly enhanced the amplitude of IpCa. Using a paired pulse protocol, the second (test) response was facilitated with inter-pulse intervals of less than 100 ms. The facilitation was greater at shorter intervals and was maximal (about 20%) at intervals of 5-10 ms. 3. When the test pulse duration was extended, the facilitation was revealed as an increased rate of IpCa Activation. From the current-Voltage relationship measured at 1 ms from onset, facilitation could be described by a shift in the half-Activation Voltage of about -4 mV. 4. IpCa facilitation was not attenuated when guanosine-5'-O-(3-thiotriphosphate) (GTPgammaS) or guanosine-5'-O-(2-thiodiphosphate) (GDPbetaS) was included in the patch pipette, suggesting that G-proteins are not involved in this phenomenon. 5. On reducing [Ca2+]o, the magnitude of facilitation diminished proportionally to the amplitude of IpCa. Replacement of [Ca2+]o by Ba2+ or Na+, or buffering of [Ca2+]i with EGTA or BAPTA attenuated IpCa facilitation. 6. We conclude that repetitive presynaptic activity can facilitate the presynaptic Ca2+ current through a Ca2+-dependent mechanism. This mechanism would be complementary to the action of residual Ca2+ on the exocytotic machinery in producing activity-dependent facilitation of synaptic responses.
Yoshinori Fujiyoshi - One of the best experts on this subject based on the ideXlab platform.
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neurosteroid pregnenolone sulfate enhances glutamatergic synaptic transmission by facilitating presynaptic calcium currents at the calyx of held of immature rats
European Journal of Neuroscience, 2006Co-Authors: Toshihide Hige, Yoshinori FujiyoshiAbstract:Pregnenolone sulfate (PREGS) is an endogenous neurosteroid widely released from neurons in the brain, and is thought to play a memory-enhancing role. At excitatory synapses PREGS facilitates transmitter release, but the underlying mechanism is not known. We addressed this issue at the calyx of Held in rat brainstem slices, where direct whole-cell recordings from giant nerve terminals are feasible. PREGS potentiated nerve-evoked excitatory postsynaptic currents (EPSCs) without affecting the amplitude of miniature EPSCs, suggesting that its site of action is presynaptic. In whole-cell recordings from calyceal nerve terminals, PREGS facilitated Ca 2+ currents, by accelerating their Activation kinetics and shifting the half-Activation Voltage toward negative potentials. PREGS had no effect on presynaptic K + currents, resting conductance or action potential waveforms. In simultaneous pre- and postsynaptic recordings, PREGS did not change the relationship between presynaptic Ca 2+ influx and EPSCs, suggesting that exocytotic machinery downstream of Ca 2+ influx is not involved in its effect. PREGS facilitated Ba 2+ currents recorded from nerve terminals and also from HEK 293 cells expressed with recombinant N- or P/Q-type Ca 2+ channels, suggesting that PREGS-induced facilitation of Voltage-gated Ca 2+ channels (VGCCs) is neither Ca 2+ dependent nor VGCC-type specific. The PREGS-induced VGCC facilitation was blocked by the PREGS scavenger (2-hydroxypropyl)-β-cyclodextrin applied from outside, but not from inside, of nerve terminals. We conclude that PREGS facilitates VGCCs in presynaptic terminals by acting from outside, thereby enhancing transmitter release. We propose that PREGS may directly modulate VGCCs acting on their extracellular domain.
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neurosteroid pregnenolone sulfate enhances glutamatergic synaptic transmission by facilitating presynaptic calcium currents at the calyx of held of immature rats
European Journal of Neuroscience, 2006Co-Authors: Toshihide Hige, Yoshinori Fujiyoshi, Tomoyuki TakahashiAbstract:Pregnenolone sulfate (PREGS) is an endogenous neurosteroid widely released from neurons in the brain, and is thought to play a memory-enhancing role. At excitatory synapses PREGS facilitates transmitter release, but the underlying mechanism is not known. We addressed this issue at the calyx of Held in rat brainstem slices, where direct whole-cell recordings from giant nerve terminals are feasible. PREGS potentiated nerve-evoked excitatory postsynaptic currents (EPSCs) without affecting the amplitude of miniature EPSCs, suggesting that its site of action is presynaptic. In whole-cell recordings from calyceal nerve terminals, PREGS facilitated Ca2+ currents, by accelerating their Activation kinetics and shifting the half-Activation Voltage toward negative potentials. PREGS had no effect on presynaptic K+ currents, resting conductance or action potential waveforms. In simultaneous pre- and postsynaptic recordings, PREGS did not change the relationship between presynaptic Ca2+ influx and EPSCs, suggesting that exocytotic machinery downstream of Ca2+ influx is not involved in its effect. PREGS facilitated Ba2+ currents recorded from nerve terminals and also from HEK 293 cells expressed with recombinant N- or P/Q-type Ca2+ channels, suggesting that PREGS-induced facilitation of Voltage-gated Ca2+ channels (VGCCs) is neither Ca2+ dependent nor VGCC-type specific. The PREGS-induced VGCC facilitation was blocked by the PREGS scavenger (2-hydroxypropyl)-beta-cyclodextrin applied from outside, but not from inside, of nerve terminals. We conclude that PREGS facilitates VGCCs in presynaptic terminals by acting from outside, thereby enhancing transmitter release. We propose that PREGS may directly modulate VGCCs acting on their extracellular domain.
