scholarly journals Ca2+-induced Ca2+ Release Phenomena in Mammalian Sympathetic Neurons Are Critically Dependent on the Rate of Rise of Trigger Ca2+

1997 ◽  
Vol 109 (2) ◽  
pp. 147-167 ◽  
Author(s):  
Arturo Hernández-Cruz ◽  
Ariel L. Escobar ◽  
Nicolás Jiménez
Keyword(s):  
Sympathetic Neurons ◽  
Feedback Gain ◽  
Release Channel ◽  
Linear Diffusion ◽  
Intracellular Release ◽  
Dependent Inactivation ◽  
Initial Release ◽  
Agonist Concentration ◽  
Fast Release

The role of ryanodine-sensitive intracellular Ca2+ stores present in nonmuscular cells is not yet completely understood. Here we examine the physiological parameters determining the dynamics of caffeine-induced Ca2+ release in individual fura-2–loaded sympathetic neurons. Two ryanodine-sensitive release components were distinguished: an early, transient release (TR) and a delayed, persistent release (PR). The TR component shows refractoriness, depends on the filling status of the store, and requires caffeine concentrations ≥10 mM. Furthermore, it is selectively suppressed by tetracaine and intracellular BAPTA, which interfere with Ca2+-mediated feedback loops, suggesting that it constitutes a Ca2+-induced Ca2+-release phenomenon. The dynamics of release is markedly affected when Sr2+ substitutes for Ca2+, indicating that Sr2+ release may operate with lower feedback gain than Ca2+ release. Our data indicate that when the initial release occurs at an adequately fast rate, Ca2+ triggers further release, producing a regenerative response, which is interrupted by depletion of releasable Ca2+ and Ca2+-dependent inactivation. A compartmentalized linear diffusion model can reproduce caffeine responses: When the Ca2+ reservoir is full, the rapid initial Ca2+ rise determines a faster occupation of the ryanodine receptor Ca2+ activation site giving rise to a regenerative release. With the store only partially loaded, the slower initial Ca2+ rise allows the inactivating site of the release channel to become occupied nearly as quickly as the activating site, thereby suppressing the initial fast release. The PR component is less dependent on the store's Ca2+ content. This study suggests that transmembrane Ca2+ influx in rat sympathetic neurons does not evoke widespread amplification by CICR because of its inability to raise [Ca2+] near the Ca2+ release channels sufficiently fast to overcome their Ca2+-dependent inactivation. Conversely, caffeine-induced Ca2+ release can undergo considerable amplification especially when Ca2+ stores are full. We propose that the primary function of ryanodine-sensitive stores in neurons and perhaps in other nonmuscular cells, is to emphasize subcellular Ca2+ gradients resulting from agonist-induced intracellular release. The amplification gain is dependent both on the agonist concentration and on the filling status of intracellular Ca2+ stores.

Ca2+ Enhances U-Type Inactivation of N-Type (CaV2.2) Calcium Current in Rat Sympathetic Neurons

2006 ◽  
Vol 96 (3) ◽  
pp. 1075-1083 ◽  
Author(s):  
Yong Sook Goo ◽  
Wonil Lim ◽  
Keith S. Elmslie
Keyword(s):  
Slow Component ◽  
Divalent Cations ◽  
Sympathetic Neurons ◽  
Slow Inactivation ◽  
Fast Inactivation ◽  
Extracellular Solution ◽  
Channel Inactivation ◽  
Dependent Inactivation ◽  
Preferential Inactivation

Ca2+-dependent inactivation (CDI) has recently been shown in heterologously expressed N-type calcium channels (CaV2.2), but CDI has been inconsistently observed in native N-current. We examined the effect of Ca2+ on N-channel inactivation in rat sympathetic neurons to determine the role of CDI on mammalian N-channels. N-current inactivated with fast (τ ∼ 150 ms) and slow (τ ∼ 3 s) components in Ba2+. Ca2+ differentially affected these components by accelerating the slow component (slow inactivation) and enhancing the amplitude of the fast component (fast inactivation). Lowering intracellular BAPTA concentration from 20 to 0.1 mM accelerated slow inactivation, but only in Ca2+ as expected from CDI. However, low BAPTA accelerated fast inactivation in either Ca2+ or Ba2+, which was unexpected. Fast inactivation was abolished with monovalent cations as the charge carrier, but slow inactivation was similar to that in Ba2+. Increased Ca2+, but not Ba2+, concentration (5–30 mM) enhanced the amplitude of fast inactivation and accelerated slow inactivation. However, the enhancement of fast inactivation was independent of Ca2+ influx, which indicates the relevant site is exposed to the extracellular solution and is inconsistent with CDI. Fast inactivation showed U-shaped voltage dependence in both Ba2+ and Ca2+, which appears to result from preferential inactivation from intermediate closed states (U-type inactivation). Taken together, the data support a role for extracellular divalent cations in modulating U-type inactivation. CDI appears to play a role in N-channel inactivation, but on a slower (sec) time scale.

scholarly journals A CACNA1A variant associated with trigeminal neuralgia alters the gating of Cav2.1 channels

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Eder Gambeta ◽  
Maria A. Gandini ◽  
Ivana A. Souza ◽  
Laurent Ferron ◽  
Gerald W. Zamponi
Keyword(s):  
Trigeminal Neuralgia ◽  
Missense Mutation ◽  
Neurotransmitter Release ◽  
Trigeminal System ◽  
Wild Type ◽  
Calcium Dependent ◽  
Whole Cell Patch Clamp ◽  
C Terminus ◽  
Dependent Inactivation

AbstractA novel missense mutation in the CACNA1A gene that encodes the pore forming α1 subunit of the CaV2.1 voltage-gated calcium channel was identified in a patient with trigeminal neuralgia. This mutation leads to a substitution of proline 2455 by histidine (P2455H) in the distal C-terminus region of the channel. Due to the well characterized role of this channel in neurotransmitter release, our aim was to characterize the biophysical properties of the P2455H variant in heterologously expressed CaV2.1 channels. Whole-cell patch clamp recordings of wild type and mutant CaV2.1 channels expressed in tsA-201 cells reveal that the mutation mediates a depolarizing shift in the voltage-dependence of activation and inactivation. Moreover, the P2455H mutant strongly reduced calcium-dependent inactivation of the channel that is consistent with an overall gain of function. Hence, the P2455H CaV2.1 missense mutation alters the gating properties of the channel, suggesting that associated changes in CaV2.1-dependent synaptic communication in the trigeminal system may contribute to the development of trigeminal neuralgia.

The neuro-cardiac junction defines an extracellular microdomain required for neurotrophic signaling

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
M Mongillo ◽  
M Franzoso ◽  
V Prando ◽  
L Dokshokova ◽  
A Di Bona ◽  
...  
Keyword(s):  
Culture Medium ◽  
Heart Diseases ◽  
Sympathetic Neurons ◽  
Confocal Imaging ◽  
Public Institution ◽  
Mdx Mice ◽  
Mouse Heart ◽  
Funding Source ◽  
Neurotrophic Signaling

Abstract Background Sympathetic neurons (SNs) innervate the myocardium with a defined topology that allows physiological modulation of cardiac activity. Neurotrophins released by cardiac cells control SN viability and myocardial distribution, which are impaired in heart diseases with reduced (e.g. heart failure) or heterogenous sympathetic stimulation (e.g. arrhythmias). We previously demonstrated that SNs interact directly with cardiomyocytes (CMs) at neuro-cardiac junctions (NCJ), and such structured contact sites allow neurons to efficiently activate β-adrenoceptors on the myocyte membrane. Aims We here asked whether NCJs are functional for retrograde (myocyte to neuron) neurotrophic signaling. Methods and results Electron microscopy and immunofluorescence on mouse heart slices and SN/CM co-cultures showed that the NGF receptor, TrkA, is preferentially found in correspondence of the NCJ. Consistently, neurons taking structured contact with CMs showed fast TrkA activation and its retrograde transport to the soma, which was monitored using live confocal imaging in cells expressing TrkA-RFP. In accord with NGF dependent effects, CM-contacted SN showed larger synaptic varicosities and did not require NGF supplementation in the culture medium. In support that NGF locally released at NCJs sustains SN viability, the neurotrophin concentration in the culture medium was 1.61 pg/mL, and did not suffice to maintain neuronal viability, which was also perturbed (66% decrease of neuronal density) by silencing NGF expression in CMs. These results support that the NCJ is essential for intercellular neurotrophin signaling. Consistently, by applying competitive inhibition of TrkA with increasing doses of K252a, we estimated NGF concentration at the contact site to be about 1000-fold higher than that released by CM in the culture medium. To seek for the structural determinants of the NCJ, we focused on dystrophin, based on the finding that the protein accumulates on the CM membrane portion contacted by SNs, as observed in mouse heart slices, and co-cultured CMs. In support of a role of CM-expressed dystrophin in neurotrophic signaling, hearts from dystrophin-KO (mdx) mice showed 74.36% decrease of innervation, with no significant changes of NGF expression. In line with the purported role of NCJs, in co-cultures between wild type SNs and mdx CMs, TrkA activation (TrkA movements toward SN soma (%): WTCM-WTSN=18±4; MDXCM-WTSN= 12±3; p<0,05) and neuronal survival were reduced. Conclusions Taken together, our results suggest that NGF-dependent signaling to SNs requires a direct and specialized interaction with myocytes, and that loss of dystrophin at the CM membrane impairs retrograde signaling to the neurons leading to cardiac sympathetic dys-innervation. Funding Acknowledgement Type of funding source: Public Institution(s). Main funding source(s): University of Padova

scholarly journals Crystal structures of Ca2+–calmodulin bound to NaV C-terminal regions suggest role for EF-hand domain in binding and inactivation

2019 ◽  
Vol 116 (22) ◽  
pp. 10763-10772 ◽  
Author(s):  
Bernd R. Gardill ◽  
Ricardo E. Rivera-Acevedo ◽  
Ching-Chieh Tung ◽  
Filip Van Petegem
Keyword(s):  
Crystal Structure ◽  
Binding Sites ◽  
Binding Mode ◽  
Conformational Switch ◽  
Channel Inactivation ◽  
Dependent Inactivation ◽  
Ef Hand ◽  
Inherited Arrhythmia ◽  
A Site

Voltage-gated sodium (NaV) and calcium channels (CaV) form targets for calmodulin (CaM), which affects channel inactivation properties. A major interaction site for CaM resides in the C-terminal (CT) region, consisting of an IQ domain downstream of an EF-hand domain. We present a crystal structure of fully Ca2+-occupied CaM, bound to the CT of NaV1.5. The structure shows that the C-terminal lobe binds to a site ∼90° rotated relative to a previous site reported for an apoCaM complex with the NaV1.5 CT and for ternary complexes containing fibroblast growth factor homologous factors (FHF). We show that the binding of FHFs forces the EF-hand domain in a conformation that does not allow binding of the Ca2+-occupied C-lobe of CaM. These observations highlight the central role of the EF-hand domain in modulating the binding mode of CaM. The binding sites for Ca2+-free and Ca2+-occupied CaM contain targets for mutations linked to long-QT syndrome, a type of inherited arrhythmia. The related NaV1.4 channel has been shown to undergo Ca2+-dependent inactivation (CDI) akin to CaVs. We present a crystal structure of Ca2+/CaM bound to the NaV1.4 IQ domain, which shows a binding mode that would clash with the EF-hand domain. We postulate the relative reorientation of the EF-hand domain and the IQ domain as a possible conformational switch that underlies CDI.

Halothane Inhibition of Recombinant Cardiac L-type Ca2+ Channels Expressed in HEK-293 Cells

2005 ◽  
Vol 103 (6) ◽  
pp. 1156-1166 ◽  
Author(s):  
Kevin J. Gingrich ◽  
Son Tran ◽  
Igor M. Nikonorov ◽  
Thomas J. Blanck
Keyword(s):  
Charge Transfer ◽  
Calcium Channels ◽  
Dependent Manner ◽  
Current Time ◽  
Hek 293 ◽  
Hek 293 Cells ◽  
293 Cells ◽  
Dependent Inactivation ◽  
Voltage Dependent

Background Volatile anesthetics depress cardiac contractility, which involves inhibition of cardiac L-type calcium channels. To explore the role of voltage-dependent inactivation, the authors analyzed halothane effects on recombinant cardiac L-type calcium channels (alpha1Cbeta2a and alpha1Cbeta2aalpha2/delta1), which differ by the alpha2/delta1 subunit and consequently voltage-dependent inactivation. Methods HEK-293 cells were transiently cotransfected with complementary DNAs encoding alpha1C tagged with green fluorescent protein and beta2a, with and without alpha2/delta1. Halothane effects on macroscopic barium currents were recorded using patch clamp methodology from cells expressing alpha1Cbeta2a and alpha1Cbeta2aalpha2/delta1 as identified by fluorescence microscopy. Results Halothane inhibited peak current (I(peak)) and enhanced apparent inactivation (reported by end pulse current amplitude of 300-ms depolarizations [I300]) in a concentration-dependent manner in both channel types. alpha2/delta1 coexpression shifted relations leftward as reported by the 50% inhibitory concentration of I(peak) and I300/I(peak)for alpha1Cbeta2a (1.8 and 14.5 mm, respectively) and alpha1Cbeta2aalpha2/delta1 (0.74 and 1.36 mm, respectively). Halothane reduced transmembrane charge transfer primarily through I(peak) depression and not by enhancement of macroscopic inactivation for both channels. Conclusions The results indicate that phenotypic features arising from alpha2/delta1 coexpression play a key role in halothane inhibition of cardiac L-type calcium channels. These features included marked effects on I(peak) inhibition, which is the principal determinant of charge transfer reductions. I(peak) depression arises primarily from transitions to nonactivatable states at resting membrane potentials. The findings point to the importance of halothane interactions with states present at resting membrane potential and discount the role of inactivation apparent in current time courses in determining transmembrane charge transfer.

Abstract P201: Negative Regulation Of PVN Pre-sympathetic Neuronal Activity By Resident Microglia

Hypertension ◽  
2021 ◽  
Vol 78 (Suppl_1) ◽  
Author(s):  
Peng Shi ◽  
Yunfan Lin ◽  
Qianqian Bi ◽  
Guo Cheng ◽  
Xiao Shen
Keyword(s):  
Neuronal Excitability ◽  
Sympathetic Neurons ◽  
Potassium Currents ◽  
Firing Frequency ◽  
Electrophysiological Recording ◽  
Recombinant Peptide ◽  
Regulatory Circuits ◽  
Brain Parenchymal ◽  
Parenchymal Cells

Hypothalamic paraventricular nucleus (PVN) is a critical integrating region in controlling peripheral sympathetic tonicity. While the vast studies have unraveled the regulatory circuits affecting PVN pre-sympathetic neurons, local factors for maintaining the homeostasis of neuronal excitability are barely understood. In the present study we investigated the role of microglia, the primary resident immune cells of the CNS, in this context. By electrophysiological recording, we found that loss of resident microglia induced an increased firing frequency and attenuated outward potassium currents in the PVN pre-sympathetic neurons, tachycardia and impaired heart rate variability. Combining the transcriptomics analysis of the PVN microglia, we identified a releasable factor, which was dominantly expressed in microglia compared to other brain parenchymal cells. ICV infusion of the recombinant peptide restored potassium currents in the PVN pre-sympathetic neurons and autonomic function in microglia-depleted mice. In summary, our results provided a novel intrinsic regulatory mechanism by which microglia suppress neuronal over excitation in physiological condition.

Tau confers drug stability but not cold stability to microtubules in living cells

1994 ◽  
Vol 107 (1) ◽  
pp. 135-143 ◽  
Author(s):  
P.W. Baas ◽  
T.P. Pienkowski ◽  
K.A. Cimbalnik ◽  
K. Toyama ◽  
S. Bakalis ◽  
...  
Keyword(s):  
Sympathetic Neurons ◽  
Drug Stability ◽  
Living Cells ◽  
Microtubule Depolymerization ◽  
Cell Biol ◽  
Cold Stability ◽  
The Stability ◽  
Cultured Sympathetic Neurons ◽  
Very High

We previously defined two classes of microtubule polymer in the axons of cultured sympathetic neurons that differ in their sensitivity to nocodazole by roughly 35-fold (Baas and Black (1990) J. Cell Biol. 111, 495–509). Here we demonstrate that virtually all of the microtubule polymer in these axons, including the drug-labile polymer, is stable to cold. What factors account for the unique stability properties of axonal microtubules? In the present study, we have focused on the role of tau, a microtubule-associated protein that is highly enriched in the axon, in determining the stability of microtubules to nocodazole and/or cold in living cells. We used a baculovirus vector to express very high levels of tau in insect ovarian Sf9 cells. The cells respond by extending processes that contain dense bundles of microtubules (Knops et al. (1991) J. Cell Biol. 114, 725–734). Cells induced to express tau were treated with either cold or 2 micrograms/ml nocodazole for times ranging from 5 minutes to 6 hours. The results with each treatment were very different from one another. Virtually all of the polymer was depolymerized within the first 30 minutes in cold, while little or no microtubule depolymerization was detected even after 6 hours in nocodazole. Based on these results, we conclude that tau is almost certainly a factor in conferring drug stability to axonal microtubules, but that factors other than or in addition to tau are required to confer cold stability.

Angiotensin II antagonists in dehydrated rabbits without baroreceptor reflexes

1977 ◽  
Vol 232 (2) ◽  
pp. H110-H113
Author(s):  
N. C. Trippodo ◽  
T. G. Coleman ◽  
A. W. Cowley ◽  
A. C. Guyton
Keyword(s):  
Blood Pressure ◽  
Angiotensin Ii ◽  
Blood Pressure Regulation ◽  
Pressure Effects ◽  
Feedback Gain ◽  
Plasma Sodium ◽  
Pressure Regulation ◽  
Angiotensin Ii Antagonists ◽  
Baroreceptor Reflexes

Blood pressure effects of angiotensin II antagonists were studied in sham-operated and baroreceptor-denervated rabbits in the normal water-replete state or after 6 days of water deprivation (dehydrated). Experiments were performed in awake rabbits. Dehydrated rabbits had significantly higher plasma sodium concentrations, hematocrits, and plasma renin activities, but lower plasma potassium concentrations and body weights than water-replete rabbits. Administration of angiotensin II antagonists caused a significant decrease in mean arterial pressure in dehydrated rabbits (-16 mmHg in sham-dehydrated and -19 mmHg in denervated-dehydrated) but not in water-replete ones, whether the baroreceptor reflexes were intact or not (-1 mmHg in sham replete and -4 mmHg in denervated replete). The open-loop feedback gain of the renin-angiotensin system in blood pressure control was calculated as -1.6. The results demonstrate an important role of angiotensin II in blood pressure regulation during the high-renin, dehydrated state, but not during the normal renin, water-replete state. Abolishment of baroreceptor reflexes did not unmask an important role of normal levels of angiotensin II in blood pressure regulation.

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