Sigma-1 receptor antagonism restores injury-induced decrease of voltage-gated Ca2+ current in sensory neurons - PubMed (original) (raw)

Sigma-1 receptor antagonism restores injury-induced decrease of voltage-gated Ca2+ current in sensory neurons

Bin Pan et al. J Pharmacol Exp Ther. 2014 Aug.

Abstract

Sigma-1 receptor (σ1R), an endoplasmic reticulum-chaperone protein, can modulate painful response after peripheral nerve injury. We have demonstrated that voltage-gated calcium current is inhibited in axotomized sensory neurons. We examined whether σ1R contributes to the sensory dysfunction of voltage-gated calcium channel (VGCC) after peripheral nerve injury through electrophysiological approach in dissociated rat dorsal root ganglion (DRG) neurons. Animals received either skin incision (Control) or spinal nerve ligation (SNL). Both σ1R agonists, (+)pentazocine (PTZ) and DTG [1,3-di-(2-tolyl)guanidine], dose dependently inhibited calcium current (ICa) with Ba(2+) as charge carrier in control sensory neurons. The inhibitory effect of σ1R agonists on ICa was blocked by σ1R antagonist, BD1063 (1-[2-(3,4-dichlorophenyl)ethyl]-4-m​ethylpiperazine dihydrochloride) or BD1047 (N-[2-(3,4-dichlorophenyl)ethyl]-N-m​ethyl-2-(dimethylamino)ethylamine dihydrobromide). PTZ and DTG showed similar effect on ICa in axotomized fifth DRG neurons (SNL L5). Both PTZ and DTG shifted the voltage-dependent activation and steady-state inactivation of VGCC to the left and accelerated VGCC inactivation rate in both Control and axotomized L5 SNL DRG neurons. The σ1R antagonist, BD1063 (10 μM), increases ICa in SNL L5 neurons but had no effect on Control and noninjured fourth lumbar neurons in SNL rats. Together, the findings suggest that activation of σR1 decreases ICa in sensory neurons and may play a pivotal role in pain generation.

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Figures

Fig. 1.

Fig. 1.

_σ_1R agonists inhibit voltage-gated calcium current in control sensory neurons. After whole-cell configuration, the _I_Ca was measured with a square wave voltage command (holding potential at −90 mV and step to 0 mV for 100 milliseconds) using Ba2+ as charge carrier. PTZ (A) reduced _I_Ca (sample traces, top panel; averaged time data, bottom panel) in a dose-related fashion (B), as did DTG (C and D). N = 4–10 for each concentration.

Fig. 2.

Fig. 2.

Effect of PTZ and DTG on _I_Ca is mediated by _σ_1R. After whole-cell configuration, the _I_Ca was measured with a square wave voltage command (holding potential at −100 mV and step to 0 mV for 50 milliseconds). Cells were preincubated with BD1047 or BD1063 for 15 minutes before PTZ and DTG administration. (A) Sample traces from control animal showed that DTG (10 _μ_M) inhibited _I_Ca and the inhibition was prevented by BD1063 (10 _μ_M). (B) Sample traces from control animal showed that PTZ (100 _μ_M) inhibited _I_Ca and the inhibition was prevented by BD1047 (10 _μ_M). Summary data of _I_Ca inhibition were derived from the current density, which was normalized to its baseline current density. Mean ± S.E.M.; number in bars represents the sample size; ***P < 0.001. BL, baseline.

Fig. 3.

Fig. 3.

_σ_1R modulates kinetic properties of VGCC in control animals. (A) _I_Ca was measured with a square wave voltage command (200 milliseconds; 10 mV increment from −100 to 50 mV; holding potential at −65 mV). Average inward current density (pA/pF) against current-voltage relationship (I-V) elicited by PTZ (50 _μ_M) and DTG (100 _μ_M) was fit to a single Boltzmann function (see Materials and Methods). (B) Voltage-dependent activation of VGCC was derived from the I-V curve by Boltzmann analysis. (C) The steady-state inactivation of _I_Ca was measured during a 4-second square wave depolarization (−100 to 30 mV in 10-mV increments) with _I_Ca determined by a subsequent test pulse (20 milliseconds to 0 mV). Results are normalized to maximal peak current (I/_I_max). (D) A simple step protocol (−100 to 0 mV for 2 seconds) was applied to measure the inactivation kinetics by a two-exponential function (_τ_1 and _τ_2). Inset figures revealed summary data of maximal conductance (_G_max; A), voltage at which current is half-maximal (_V_1/2; B and C), and inactivation constant (τ; D). Mean ± S.E.M.; number in bars represents the sample size; *P < 0.05; **P < 0.01; ***P < 0.001. BL, baseline.

Fig. 4.

Fig. 4.

Inhibition of _σ_1R agonists on _I_Ca encompasses multiple _I_Ca subtypes in control sensory neurons. (A) In whole-cell configuration, _I_Ca was measured with a square wave voltage command (holding potential at −90 mV and step to 0 mV for 150 milliseconds). Sample current traces (top) and averaged data (bottom panel) show response to _σ_1R activation by PTZ (100 _µ_M) after blockade of N-type channels with _ω_-conotoxin GVIA (GVIA, 200 nM). (B) Similarly, sensitivity to PTZ is demonstrated after blockade of L-type channels with nimodipine (5 _µ_M). (C) T-type _I_Ca was triggered by depolarizations from a holding potential at −100 mV and step to −30 mV after incubation (20 minutes) with R-type current blocker SNX-482 (200 nM) and P/Q-type current blocker _ω_-conotoxin MVIIC (200 nM), and addition of L-type current blocker nimodipine (5 _µ_M) and GVIA (200 nM) to the recording bath. These currents also showed sensitivity to PTZ. (D) Summary data. Mean ± S.E.M.; number in bars represents the sample size; **P < 0.01; ***P < 0.001. BL, baseline.

Fig. 5.

Fig. 5.

_σ_1R modulates kinetics of VGCC in axotomized animals. After whole-cell configuration, the current-voltage relationship (I-V), voltage-dependent activation, and steady-state inactivation of the _I_Ca were measured as described in Fig. 4 using Ba2+ as charge carrier in SNL L5 DRG sensory neurons. (A) Average inward current density (pA/pF) against I-V relationship elicited by PTZ (50 _μ_M) and DTG (100 _μ_M) was fit to a single Boltzmann function (see Materials and Methods). (B) Voltage-dependent activation of VGCC was derived from the I-V curve. (C) In steady-state inactivation, _V_1/2 was shifted toward a hyperpolarization direction. (D) Sample traces showed that both PTZ and DTG administration decreased slow inactivation constant (_τ_2). Inset figures revealed summary data of maximal conductance (_G_max; A), voltage at which current is half-maximal (_V_1/2; B and C), and inactivation constant (τ; D). Mean ± S.E.M.; number in bars represents the sample size; *P < 0.05; **P < 0.01; ***P < 0.001. BL, baseline.

Fig. 6.

Fig. 6.

_σ_1R antagonist, BD1063 (10 _μ_M), increased _I_Ca in axotomized fifth (SNL L5) DRG sensory neurons. After whole-cell configuration, the _I_Ca was measured with a square wave voltage command (holding potential at −100 mV and step to 0 mV for 20 milliseconds) using Ba2+ as charge carrier. Sample current traces (A and B) showed that BD1063 (10 _μ_M) did not affect _I_Ca in control and noninjured neighboring fourth (SNL L4) DRG neurons. (C) BD1063 administration increased the _I_Ca in SNL L5 neurons. Summary data demonstrated that application of BD1063 (10 _μ_M) was able to increase in SNL L5 but not control and SNL L4 neurons. Mean ± S.E.M.; number in bars represents the sample size; **P < 0.01. BL, baseline.

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