Sildenafil and Neuropathic Pain/Hemodynamics It has been suggested that cGMP is involved in antinoci- ception.8 In line with this observation, dibutyryl-cGMP and 8-bromo-cGMP showed antinociception in a modification of the Randall-Selitto hyperalgesia model and in a neuro- pathic pain model, respectively.7,9 Furthermore, sildenafil, a phosphodiesterase 5 inhibitor, caused antinociception in carrageenan-induced hyperalgesia, the writhing test and the formalin test.10-14 In a previous study, increasing cGMP levels with the phosphodiesterase inhibitor zaprinast enha- nced the γ-aminobutyric acid (GABA) release.15 Recently, the antinociception of sildenafil was shown to be reversed by a GABAB receptor antagonist in the formalin test.16 These observations suggest that GABA receptors may be involved in the activity of sildenafil. On the other hand, the activity of phosphodiesterase 5 was detected in smooth muscle cells of vessels.17 Also, cGMP plays important role in the regulation of vascular tone,18 and it has recently been reported that phosphodies- terase 5 is the key enzyme involved in the regulation of the cGMP-associated vascular relaxation.19 Therefore, hemo- dynamic change due to vasodilating effect of phosphodie- sterase 5 inhibitor is expected. In this study, we examined the effect of intravenous sildenafil in a rat neuropathic pain model and attempted to clarify the role of GABA receptors in the action of sildenafil. Furthermore, we observed the change of hemodynamics following administration of intravenous sildenafil in heal- thy rats. MATERIALS AND METHODS Animal preparation This study proposal was reviewed and approved by the Institutional Animal Care Committee, Research Institute of Medical Science, Chonnam National University. Sprague- Dawley male rats, weighing 100-200 g, were used in all experiments. Animals were acclimated to the laboratory environment for 5-7 days before being used in the study. While in the home cage environment, the animals were allowed free access to a standard rat diet and tap water. Room temperature was maintained at 20-23˚C with 12 : 12 h light/dark cycle. Neuropathic pain model Neuropathic pain was evoked by spinal nerve ligation of experimental rats as previously described.20 Briefly, the left L5 and L6 spinal nerves of rats were isolated adjacent to the vertebral column during isoflurane anesthesia and tightly ligated with a 6-0 silk suture distal to the dorsal root ganglia. Care was taken to avoid injury of the L4 spinal nerve. Following surgery, development of neuropathic pain was evaluated daily by measuring the mechanical sensitivity of the injured paw. Animals were considered to be in neuro- pathic pain when they exhibited mechanical allodynia i.e., paw flinching behavior response to the application of a bending force of less than 4 g. All animals were allowed to recover for at least 1 week. Assessment of mechanical allodynia Mechanical allodynia was measured using calibrated von Frey filaments as previously described.21 Rats were placed into inverted individual plastic containers (20×12.5×20 cm) on top of a suspended wire mesh grid, and acclimated to the test chambers for 20 min. A series of eight von Frey filaments (0.4, 0.7, 1.2, 2.0, 3.6, 5.5, 8.5, and 15 g) were applied vertically to the plantar surface of the hindpaw for 5 s while the hair was bent. Brisk withdrawal or paw flinching was considered positive responses. In the absence of a response at a pressure of 15 g, animals were assigned to this cut-off value. Tests were performed in duplicate with an approximate 3 min test-free period between with- drawal responses, and their average was used. Positive res- ponses included an abrupt withdrawal of the hind paw from the stimulus, or flinching behavior immediately following removal of the stimulus. Drugs and administration The following drugs were used in this study: sildenafil (phosphodiesterase 5 inhibitor), bicuculline (GABAA receptor antagonist, Sigma Aldrich Co., St. Louis, MO, USA) and saclofen (GABAB receptor antagonist, Sigma). Sildenafil was kindly provided by Korea Pfizer. All drugs were dissolved in normal saline. For the intravenous admi- nistration of these agents, the tail vein was used. For intra- venous administration, drugs and saline were injected in volumes of 3 mL/kg. Hemodynamic measurement In order to measure hemodynamic changes, a polyethy- lene-50 catheter was inserted into the tail artery under isoflurane (3-4%)/O2 anesthesia, and the rats were then restrained in a restraint cylinder. The catheter was flushed with 0.5 mL of heparinized saline. The arterial line was connected to a pressure transducer of monitor (Datex- Ohmeda AS/3, GE Healthcare, Helsinki, Finland) for continuous recording of blood pressure and heart rate. Experimental paradigm Seven days after nerve ligation, sildenafil was administered intravenously and mechanical threshold for paw flinching was measured at 15, 30, 45, 60, 90, 120, 150, and 180 min after delivery of sildenafil. Measurement of the mechanical threshold for paw flinching was also carried out before Yonsei Med J http://www.eymj.org Volume 51 Number 1 January 2010 83