A novel proline-rich M-superfamily conotoxin that can simultaneously affect sodium, potassium and calcium currents

Background Conotoxins have become a research hotspot in the neuropharmacology field for their high activity and specificity in targeting ion channels and neurotransmitter receptors. There have been reports of a conotoxin acting on two ion channels, but rare reports of a conotoxin acting on three ion channels. Methods Vr3a, a proline-rich M-superfamily conotoxin from a worm-hunting Conus varius, was obtained by solid-phase synthesis and identified by mass spectrometry. The effects of synthesized Vr3a on sodium, potassium and calcium currents were tested on rat DRG cells by patch clamp experiments. The further effects of Vr3a on human Cav1.2 and Cav2.2 currents were tested on HEK293 cells. Results About 10 μM Vr3a has no effects on the peak sodium currents, but can induce a ~10 mV shift in a polarizing direction in the current-voltage relationship. In addition, 10 μM Vr3a can increase 19.61 ± 5.12% of the peak potassium currents and do not induce a shift in the current-voltage relationship. An amount of 10 μM Vr3a can inhibit 31.26% ± 4.53% of the peak calcium currents and do not induce a shift in the current-voltage relationship. The IC50 value of Vr3a on calcium channel currents in rat DRG neurons is 19.28 ± 4.32 μM. Moreover, 10 μM Vr3a can inhibit 15.32% ± 5.41% of the human Cav1.2 currents and 12.86% ± 4.93% of the human Cav2.2 currents. Conclusions Vr3a can simultaneously affect sodium, potassium and calcium currents. This novel triple-target conotoxin Vr3a expands understanding of conotoxin functions.(AU)

Analgesic effect of intrathecal SO3 on rats with chronic constriction injury and its regulation of intracellular calcium ion concentration of spinal dorsal root ganglion neurons / 中国药理学通报

Aim To evaluate the effect of intrathecal administration of ?-conopeptide SO3,an neotype N-type calcium channel blocker,on pain related behaviors and changes in intracellular calcium ion concentration of the DRG neurons in a rat model of chronic constriction injury(CCI).Methods 40 male SD rats weighing 230~270 g were randomly divided into five groups of 8 animals each.Rats in the N group served as unoperated controls;in group C the rats were placed four loose ligatures around the right sciatic nerve;in group CN 20?l normal Saline was administered 14 days after operation;in group CS1 SO3 600 ng was administered the same time as group CN;in group CS7 SO3(30 ng?h~(-1)) was chronically administered for 7 days from the 7th day post-operation.Thermal hyperalgesia and mechanical allodynia were tested by measuring paw withdrawal latency(PWL) in response to thermal stimulation and by von-Frey filaments.The animals were decapitated on time.The DRGs(L_4-L_6) were homogenized into single cells and intracellular calcium ion concentration([Ca~(2+)]_i) was measured by flow cytometry.Results The CCI procedure leads to the development of spontaneous pain,heat hyperalgesia and tactile allodynia in the hind foot innervated by the injured nerve.Intrathecal of SO3 produced significant antinociceptive effects on both thermal hyperalgesia and mechanical allodynia.In CCI rats,significant increases were observed in [Ca~(2+)]_i on the contralateral side and ipsilateral side of DRGs on day 14 after operation.I.t NS did not show any difference in [Ca~(2+)]_i to those in group C.The treatment with i.t SO3 600 ng suppressed the increases in [Ca~(2+)]_i of bilateral side of DRGs.Chronically administration of SO3 suppressed the increases in [Ca~(2+)]_i of bilateral side of DRGs further.Conclusion Selective N-type VSCC blockers are potent and efficacious antinociceptive agents when they are administered by the spinal route,and they are effective in a rat model of neuropathic pain.The antinociceptive effect provided by intrathecal SO3 is associated with N-type calcium channel,which suppresses the increases in [Ca~(2+)]_i in the DRGs.