Selectivity of ω-CgTx-MVIIC toxin from Conus magus on calcium currents in enteric neurons

doi:10.1016/S0024-3205(99)00213-1

Life Sciences

Volume 64, Issue 26, 21 May 1999, Pages PL305-PL310

https://doi.org/10.1016/S0024-3205(99)00213-1 Get rights and content

Abstract

Whole-cell perforated patch clamp recordings were used to analyze selectivity of ω-CgTx-MVIIC toxin for voltage-dependent calcium currents in cultured myenteric neurons from guinea-pig small intestine. ω-CgTx-MVIIC (300 nM) blocked 37 ± 9% of the peak current activated from −80 mV in 15 neurons by mostly affecting the plateau phase of the current. The toxin suppressed peak current activated from −30 mV dose-dependently with an ICso of 70 ± 8 nM. The blockade was complete at toxin concentrations of 1 μM. Thus, it appears that ω-CgTx-MVIIC blocks high voltage activated (HVA) calcium channels in the myenteric neurons unselectively as well as other types of HVA Ca²⁺ channels including P and Q channels.

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Cited by (21)

Pharmacological characterisation of voltage-dependent Ca<sup>2+</sup> channels in isolated ganglia from the myenteric plexus
2005, Life Sciences
Voltage-dependent Ca²⁺ channels of fura-2-loaded ganglionic cells from the myenteric plexus of newborn rats were pharmacologically characterised. In contrast to completely dissociated myenteric cells, intact ganglia showed a stronger loading with the Ca²⁺-sensitive dye and a reproducible stimulation of the fura-2 signal by the cholinergic agonist, carbachol. A depolarisation-induced increase in the intracellular Ca²⁺ concentration ([Ca²⁺]_i) was induced by superfusion with 35 mmol l⁻ ¹ KCl. This increase in [Ca²⁺]_i was sensitive to Ni²⁺ and Co²⁺ as well as ω-conotoxin MVIIA, ω-agatoxin IVA, and SNX-482. The strongest inhibition was achieved by nifedipine (5 × 10⁻ ⁷ mol l⁻ ¹) and ω-conotoxin GVIA (4.3 × 10⁻ ⁷ mol l⁻ ¹). These two blockers also inhibited the [Ca²⁺]_i increase evoked by nicotinic receptor stimulation. Consequently, isolated myenteric ganglia in culture express different types of voltage-dependent Ca²⁺ channels, from which the L- and the N-type seem to be the most important. When exposed to mediators of inflammation such as tumor necrosis factor-α (TNF-α) or different prostaglandins, no pronounced alterations in the fura-2 ratio were observed suggesting that changes in the Ca²⁺-signalling are not centrally involved in the response of enteric ganglionic cells to these paracrine substances.
Calcium channels in enteric neurons
2003, Current Opinion in Pharmacology
The enteric nervous system consists of two ganglionated neural networks within the gut wall that contain as many neurons as the spinal cord. Connections exist between the neurons in these two networks, enabling motility and secretion to be coordinated. It is becoming increasingly apparent that Ca²⁺ movements across the cell membrane and between various intracellular compartments play a major role in the regulation of neuronal excitability and neurotransmitter release.
Action potential afterdepolarization mediated by a Ca<sup>2+</sup>-activated cation conductance in myenteric AH neurons
2002, Neuroscience
We investigated the nature of afterdepolarizing potentials in AH neurons from the guinea-pig duodenum using whole-cell patch-clamp recordings in intact myenteric ganglia. Afterdepolarizing potentials were minimally activated following action-potential firing under normal conditions, but after application of charybdotoxin (40 nM) or tetraethyl ammonium (TEA; 10–20 mM) to the bathing solution, prominent afterdepolarizing potentials followed action potentials. The whole-cell current underlying afterdepolarizing potentials (I_ADP) in the presence of TEA (10–20 mM) reversed at −38 mV and was not voltage-dependent. Reduction of NaCl in the bathing (Krebs) solution to 58 mM shifted the reversal potential of the I_ADP to −58 mV, suggesting that the current underlying the afterdepolarizing potential was carried by a mixture of cations. The relative contributions of Na⁺ and K⁺ to this current were estimated to be about 1:5. Substitution of external Na⁺ with N-methyl D-glucamine blocked the current while replacement of internal Cl⁻ with gluconate did not block the I_ADP. The I_ADP was also inhibited when CsCl-filled patch pipettes were used. The I_ADP was blocked or substantially decreased in amplitude in the presence of N-type Ca²⁺ channel antagonists, ω-conotoxin GVIA and ω-conotoxin MVIIC, respectively, and was eliminated by external Cd²⁺, indicating that it was dependent on Ca²⁺ entry. The I_ADP was also inhibited by ryanodine (10–20 μM), indicating that Ca²⁺-induced Ca²⁺ release was involved in its activation. Niflumic acid consistently inhibited the I_ADP with an IC₅₀ of 63 μM. Using antibodies against the pore-forming subunits of L-, N- and P/Q-type voltage-gated Ca²⁺ channels, we have demonstrated that myenteric AH neurons express N- and P/Q, but not L-type voltage-gated Ca²⁺ channels. We conclude that the ADP in myenteric AH neurons, in the presence of an L-type Ca²⁺-channel blocker, is generated by the opening of Ca²⁺-activated non-selective cation channels following action potential-mediated Ca²⁺ entry mainly through N-type Ca²⁺ channels. Ca²⁺ release from ryanodine-sensitive stores triggered by Ca²⁺ entry contributes significantly to the activation of this current.
Sodium conductance in cultured myenteric AH-type neurons from guinea-pig small intestine
2002, Autonomic Neuroscience: Basic and Clinical
Whole-cell patch clamp methods were used to investigate sodium conductance in after-hyperpolarization-type (AH) enteric neurons in culture after dissociation from the myenteric plexus of guinea-pig small intestine. Inward current carried by Na⁺ (I_Na) was identified and its current–voltage characteristics were compared with those for inward Ca²⁺ current (I_Ca). The I_Na current was a rapidly inactivating current relative to I_Ca. Application of tetrodotoxin (TTX) blocked I_Na with an EC₅₀ of 10.7 nM. Activation curves for I_Na showed a rapid decrease in time to peak for test potentials from holding potentials of −80 mV to between −40 and −10 mV. Voltage-dependence of steady-state inactivation curves for I_Na was fit to the Boltzmann equation with potential for half-inactivation (V_1/2)=−55.6 mV and slope factor (k)=6.4 mV. Steady-state inactivation for I_Ca fit the Boltzmann equation with a V_1/2=−38.9 mV and k=14.4 mV. Kinetics for inactivation of I_Na were voltage dependent at potentials between −70 and −30 mV and accelerated and became less voltage-dependent at more positive potentials. The time constant (τ) for inactivation at −70 mV was τ=161±23 ms and decreased to τ=2.3±0.2 ms at −30 mV. Rapid acceleration of inactivation occurred between −50 and −40 mV. This was also the range where activation began. Recovery from inactivation with the membrane potential clamped at −100 or −80 mV was rapid and fit by a single exponential with τ=7.3±1.1 ms for −100 mV and 21.5±5.1 ms for −80 mV. The results suggest that AH-type enteric neurons have only one type of Na⁺ channel that behaves like the “classical” voltage-gated tetrodotoxin-sensitive fast channel. The findings support the hypothesis that I_Na current is an important factor in determination of excitability and firing behavior in AH neurons. I_Na and I_Ca together determine the properties of the rising phase of the spike and thereby contribute to global determinants of excitability as the neurons are exposed to multiple depolarizing and hyperpolarizing stimuli from synaptic inputs and mediators released from enteroparacrine cells.
Calcium channels coupled to depolarization-evoked glutamate release in the myenteric plexus of guinea-pig ileum
2000, Neuroscience
Glutamate is the major excitatory neurotransmitter in the CNS. The recent characterization of glutamate as a neurotransmitter in the enteric nervous system opened a new line of investigation concerning the role of glutamate in that system. The present study aimed to further characterize the enteric glutamate release and the calcium channels coupled to it. For this study the myenteric plexus–longitudinal muscle of guinea-pig ileum was stimulated with potassium chloride or with electrical pulses. The released glutamate was detected by spectrofluorimetry. Laser scanning confocal microscopy was used for analysis of immunolabeled enteric tissue for co-localization studies of calcium channels (N- and P/Q-type) and glutamate transporters (EAAC1).
Here we report the effects of known Ca²⁺-channel blockers on glutamate release evoked by KCl-depolarization or electrical stimulation in the myenteric plexus. We find that N-type Ca²⁺ channels control a major portion of evoked glutamate release from this system, with a very small contribution from L-type Ca²⁺ channels. Moreover, α_1A-like (P-type Ca²⁺ channel) and α_1B-like (N-type Ca²⁺ channel) immunoreactivity co-localized with glutamate transporters in the myenteric plexus. In addition, KCl-evoked or electrically stimulated glutamate release was sensitive to ω-agatoxin IVA, in a frequency-dependent manner, suggesting that P-type channels are also coupled to the release of glutamate. We, thus, conclude that both N-type and P-type Ca²⁺ channels control most of the evoked glutamate release from the enteric nervous sytem, as also occurs in some parts of the CNS.
VPAC receptor subtypes tune purinergic Neuron-to-Glia communication in the Murine submucosal plexus
2017, Frontiers in Cellular Neuroscience

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Pharmacology letter Accelerated communicationSelectivity of ω-CgTx-MVIIC toxin from Conus magus on calcium currents in enteric neurons

Abstract

Pharmac.Ther.

Neuropharmacology

Neuron (Lond)

Eur. J.Pharmacol.

Neuropharmacology

Neuroscience

Gastroenterology

Br.J.Pharmacol.

Nature (Lond)

Nature (Lond)

Pharmacology letter Accelerated communication
Selectivity of ω-CgTx-MVIIC toxin from Conus magus on calcium currents in enteric neurons