Inhibitory Effect Of ZD7288 On Sensitivity In Rats With Visceral Pain And Its Mechanism

ObjectiveThe effect of ZD7288 on sensitivity of rats in acute and chronic visceral pain were observed and the possible mechanism was investigated in order to deeply understand the pathogenesy of visceral pain,which could supply target point to the pharmacotherapy of visceral pain..Methods①Adult male Sprague-Dawley(SD)rats were used as study subject.Acetic acid was injected to the colons of rats to establish the model of acute visceral pain. Abdominal withdrawal reflex(AWR)and amplitudes of external oblique muscle of abdomen (EOMA) were tested to assess the sensitivity of rats . ZD7288 at the dose of 50～100nM was given by intrathecal application to the model rats and the effect of ZD7288 in different dosees on the reaction to pain were observed in model rats; The expression and distribution of HCN2 channel protein in dorsal root ganglion and spinal cord were detected by immuno-histochemical techniques in control and model rats.②Neonatal male Sprague-Dawley(SD)rats were used as study subject. Model rats were received 60 mmHg colorectal distensions (CRD) once daily on post neonatal days 8-15. AWR and amplitudes of EOMA were tested to assess the sensitivity of rats when they were adult. AWR and amplitudes of EOMA were tested to assess the sensitivity of rats when they were adult . ZD7288 at the dose of 25～100nM was given by intrathecal application to the model rats and the effect of ZD7288 in different dosees on the reaction to pain were observed in model rats ; The expression and distribution of HCN2 channel protein in dorsal root ganglion and spinal cord were detected by immuno-histochemical techniques in control and model rats .Results1 Inhibitory effect of ZD7288 on sensitivity in rats with acute visceral pain and its mechanism 1.1 Character of acute inflammatory visceral pain①The scores of AWR under 20～40 mmHg CRD were significantly higher in acute model rats than those in control rats.②Compared with control rats, the pain threshold in acute model rats was significantly decreased.③Compared with control rats, the amplitudes of EOMA in acute model rats were significantly increased under 20～80 mmHg CRD.④Distal colon descendent topographic histology had inflammatory change by naked eyes and microscope in acute model rats.1.2 Inhibitory effect of ZD7288 by intrathecal application on sensitivity in rats with acute visceral pain①Compared with rats that were given saline by intrathecal application , the scores of AWR were decreased under 20～40 mmHg CRD in rats that were given 50 nM ZD7288 and under 20～80 mmHg CRD in rats that were given 100nM ZD7288;There were statistical significance in the scores of AWR under 20～80 mmHg CRD between the two groups of administration.②The pain threshold in acute model rats was increased by intrathecal application of ZD7288 in a dose-dependent manner.③Compared with rats that were given saline by intrathecal application ,the amplitudes of EOMA were decreased under 20～60 mmHg CRD in rats that were given 50nM ZD7288 and under 20～80 mmHg CRD in rats that were given 100nM ZD7288;There were statistical significance in the amplitudes of EOMA under 40～80 mmHg CRD between the two groups of administration..1.3 Morphologic study of the relationship between HCN channel and sensitivity in rats with acute visceral painExpression of HCN2 channel protein was observed in dorsal root ganglion and thoracolumbar and lunbosacral spinal cord.There were positive cells at above-mentioned parts in control and acute model rats,mainly in cell membrane and cytoplasm. The results by image analysis indicated that the expression of HCN2 channel protein at each part above-mentioned in acute model rats was increased than that in control rats.2 Inhibitory effect of ZD7288 on sensitivity in rats with chronic visceral pain and its mechanism 2. 1 Character of chronic visceral pain①The scores of AWR under 20～60 mmHg CRD were significantly higher in chronic model rats than those in control rats.②Compared with control rats, the pain threshold in chronic model rats was significantly decreased.③Compared with control rats, the amplitudes of EOMA in chronic model rats were significantly increased under 20～80 mmHg CRD.④Distal colon descendent topographic histology had no apparent change by naked eyes and microscope in control and chronic model rats.2.2 Inhibitory effect of ZD7288 by intrathecal application on sensitivity in rats with chronic visceral pain①Compared with rats that were given 25nM ZD7288 by intrathecal application , the scores of AWR were decreased under 20～40 mmHg CRD in rats that were given 50nM ZD7288 and under 20～80 mmHg CRD in rats that were given 100nM ZD7288;Compared with rats that were given 50nM ZD7288,the scores of AWR were decreased under 60～80 mmHg CRD in rats that were given 100nM ZD7288.②The pain threshold in chornic model rats was increased by intrathecal application of ZD7288 in a dose-dependent manner.③Compared with rats that were given 25nM ZD7288 by intrathecal application ,the amplitudes of EOMA were decreased under 20～80 mmHg CRD in rats that were given 50nM and 100nM ZD7288;Compared with rats that were given 50nM ZD7288,the amplitudes of EOMA were decreased under 60～80 mmHg CRD in rats that were given 100nM ZD7288 .2.3 Morphologic study of the relationship between HCN channel and sensitivity in rats with chronic visceral painExpression of HCN2 channel protein was observed and in dorsal root ganglion and thoracolumbar and lunbosacral spinal cord .There were positive cells at above-mentioned parts in control and chronic model rats,mainly in cell membrane and cytoplasm. The results by image analysis indicated that the expression of HCN2 channel protein at each part above-mentioned in chronic model rats was increased than that in control rats. Conclusion1.The reaction to pain in rats with acute visceral pain can be inhibited and the pain threshold can be increased by intrathecal application of ZD7288 in a dose-dependent manner.2.The expression of HCN2 channel protein are increased in dorsal root ganglion and in thoracolumbar and lunbosacral spinal cord in acute model rats , these suggest that HCN2 subunit might be related to the mechanism of peripheral and central sensitization of acute visceral pain.3. The reaction to pain in rats with chronic visceral pain can be inhibited and the pain threshold can be increased by intrathecal application of ZD7288 in a dose-dependent manner.4.The expression of HCN2 channel protein are increased in dorsal root ganglion and in thoracolumbar and lunbosacral spinal cord in chronic model rats, these suggest that HCN2 subunit might be related to the mechanism of peripheral and central sensitization of chronic visceral pain.