We investigated diode laser (980 nm) evoked activation of transient receptor potential proteins (TRPV1 and TRPV2). C and A-delta (A&dgr;) nociceptor families are primarily responsible for pain mediation in the peripheral nervous system. TRPV1 proteins have been associated with heat evoked pain in C fibers while A&dgr; fibers have been associated with TRPV2. Diode laser stimulation allows a margin of safety between non-invasive activation and damage 19, 22, 34. Laser pulses (20-50 ms, 0.1-10 W, 980 nm) were used to stimulate: A) in vitro: excised patches from HEK293 cells expressing TRPV1; B) in vitro: rat DRG nociceptors expressing either TRPV1 or TRPV2; and C) in vivo: C-fibers of the rat saphenous nerve (SN) trunk. Cell currents were recorded using standard patch clamp methods. The SN was also stimulated electrically with bipolar electrodes. Stimulation (20-50 ms) of HEK and DRG cells expressing TRPV1 was highly reproducible. Activation and peak currents were achieved at estimated peak temperatures of 55°C and 70°C. Threshold activation was also observed in DRG neurons expressing TRPV2. The conduction velocity for laser-activated saphenous nerve afferents was in the C fiber range (0.5-1 m/s). Electrically stimulated nerve contained stimulation artifacts and complex neural components with conduction velocities ranging from 0.3-30 m/s. Diode laser activation of TRPV1 protein is a reproducible and effective means to probe TRP activity in both in vivo and in vitro preparations
We assessed the capacity of ultrashort E-fields to activate rat cutaneous nociceptors. Experiments were
conducted in vitro on nociceptive neurons representing hairy skin and glabrous skin. Electrical and optical
recording methods were used to assess action potentials and membrane damage thresholds. Strength duration
(SD) curves were formed for E-field pulses from 500 μsec to 350 ns. There were no differences in the SD time
contant (taue (59 μsec) or ultrashort thresholds (129 V/cm at 350 ns) for hairy or glabrous skin nociceptors, for
nociceptors with distinct geometry or for nociceptors expressing different combinations of voltage sensitive
Na+ channels (TTXs and TTXr Nav) or hyperpolarization activated channels (HCN; IH). Subthreshold
activation was possible with high frequency pulsing at ultrashort durations (350 ns; 4,000 Hz). Relative to
single pulse thresholds, activation threshold could be reduced over 50% by high frequency burst trains (4,000
Hz; 1-40 msec). Nociceptors were not damaged by E-field activation. Irreversible membrane disruption
occurred at significantly higher field strength and varied by cell radius (3,266-4,240 V/cm, 350 ns, 40 Hz, 5
sec). Pulse frequency had no influence on acute membrane failure (10, 20, 40, 4,000 Hz; 5 sec).
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