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Expedited Pain Drug Discovery 

Methods and Equipment

by Dr. Michael Nemenov

Fig. 1. Diode laser pulses induce ramp-shaped heat stimuli. Time courses of diode laser induced temperature changes were recorded with open patch pipettes.

Each trace was averaged from two single measurements, filtered with an eight-pole Bessel filter (50 Hz) and a notch filter (center frequency, 50 Hz; width, 10 Hz). (A) Peak temperature was always reached at the end of the laser pulse, and was a function of pulse duration (power: 11 W). (B) Peak temperature was also a function of laser power (duration: 400 ms). (C) Calibration diagram for one pipette (mean ^ SEM), based on the average temperature coefficient of 20.14 ^ 0.018C pA21 determined in a previous study (Schwarz et al., 2000). For any given pulse duration, peak temperature was proportional to the applied laser power (W 400 ms; A 240 ms; K 120 ms; L 80 ms). Note that either 11 Wfor 240 ms or 7.3 Wfor 400 ms led to nearly the same peak temperature. (D) By simultaneously changing power and duration of the laser pulse, the same peak temperature was achieved with different slopes.

Inward currents in primary nociceptive neurons of the rat and pain sensations in humans elicited by infrared diode laser pulses.

Wolfgang Greffratha, Michael I. Nemenovb, Stefan Schwarza, Ulf Baumga¨rtnera, Hagen Vogela,

Lars Arendt-Nielsenc, Rolf-Detlef Treedea,*

aInstitute of Physiology and Pathophysiology, Johannes Gutenberg University , Saarstrasse 21, D-55099 Mainz, Germany

bLaser Medical Center, I.P. Pavlov State Medical University, Lev Tolstoi Street 6/8, St. Petersburg 197022-1, Russia

cCenter for Sensory–Motor Interaction, Aalborg University, Fredrik Bajers Vej 7, D3, DK-9220 Aalborg, Denmark

Radiant heat is often used to study nociception in vivo. We now used infrared radiation generated by a diode laser stimulator (wavelength 980 nm) to investigate transduction mechanisms for noxious heat stimuli in acutely dissociated dorsal root ganglion (DRG) neurons of rats in vitro. The laser stimulator offered the unique opportunity to test whether the same stimuli also elicit pain sensations in humans.A specific heatinduced current (Iheat) was elicited in six of 13 small DRGneurons (diameter#30 mm) tested in the whole-cell configuration of the patch–clamp mode. Current responses in the seven heat-insensitive neurons were within the range explainable by the temperature dependence of the recording setup. Iheat was characterized by: (1) non-linearity of its amplitude during a suprathreshold heat ramp as well as with stimuli of increasing intensity with an estimated threshold of 42 ^ 18C; (2) fast rise time and even faster decay time (t1=2 ¼ 96:5 ^ 5:9 and 27.7 ^ 1.5 ms, respectively); and (3) rate dependence of its induction. All three heat-sensitive neurons tested were also sensitive to capsaicin. The mean threshold for the induction of Iheat was 2.8 ^ 0.3 J mm22. The threshold for the induction of action potentials by depolarizing current pulses was significantly reduced after laser stimulation, suggesting a sensitization at the transformation stage. No such change was seen in heat-insensitive neurons that underwent the same heat stimuli. The same diode laser elicited pain sensations and laser-evoked potentials in human subjects. No significant differences were seen between the pain thresholds in hairy and in glabrous skin, probably due to the deep penetration of this laser radiation. The mean pain threshold for stimuli $200 ms in humans was 2.5 ^ 0.2 J mm22 (n ¼ 11), and did not differ from the thresholds for the induction of Iheat in vitro. Our results indicate that Iheat in primary sensory neurons can be activated by infrared laser pulses that are painful in humans. q 2002 Published by Elsevier Science B.V. on behalf of International Association for the Study of Pain. 

Infrared diode laser pulses that are painful for human subjects, activate an inward current in primary nociceptive neurons in vitro that shares many properties with Iheat that has been studied with application of heated solutions. As Iheat is likely mediated by opening of the vanilloid receptor channel VR1, brief heat stimuli generated by infrared lasers offer the opportunity to study the functional integrity of VR1-expressing nerve fibers in humans. The LEP data in humans showed a latency that corresponded to activation of myelinated afferents. This observation suggests that the diode laser pulses activated type II A-fiber mechano-heat nociceptors, a class of polymodal nociceptive afferents that are thought to express the fast Iheat and VR1. Diode lasers are adaequate as non-contact heat stimulators for human pain research, which rapidly activate capsaicin-sensitive nociceptive afferents relatively independent of their depth of termination within the skin.