Evidence that activation of 5-HT2A not 5-HT2C receptors excites the external urethral sphincter in anaesthetized female rats

Yvonne Mbaki, Andrew Ramage

UCL, London, UK

5-HT2 agonists cause inhibition of micturition by activation of central 2C receptors, and increases in urethral and bladder tone by activation of peripheral 2B & 2A receptors, respectively. The ability of 5 HT2 agonists to evoke an increase in external urethral sphincter (EUS) EMG could only be attributed to activation of either spinal 2A or 2C receptors (Mbaki and Ramage, 2008). The present experiments were designed to determine which of these subtypes caused EUS-EMG activation by giving the 5-HT2 ligands by infusion (i.v.) instead of by bolus injection, to allow a gradual increase in the plasma concentration. The 5 HT2 agonist Ro 60-0175 was infused in the absence or presence of infusions of either the 5-HT2C antagonist SB 242084 or the 5-HT2A antagonist MDL 100907 (MDL).

Experiments were performed on spontaneously breathing female Sprague-Dawley rats (250-300g) anaesthetized with isoflurane (5% in 100% oxygen) and maintained with urethane (1.2 g kg-1, i.v.). Simultaneous recordings of EUS-EMG activity, urethral, bladder and blood pressures were made. Micturition reflexes were evoked by infusion (0.1 ml min-1) of saline into the bladder. The compounds or vehicles (saline and antagonist vehicle CTE (4% in saline of a mixture of 50% cremaphor, 40% tetraglycol and 10% ethanol)) were infused for 90 min. Changes were compared with vehicle controls for baseline EUS-EMG activity and urethral pressure by two-way ANOVA and the micturition reflex by unpaired Student’s t test. All values are expressed as mean±s.e.mean. P < 0.05 was considered to be significant.

Infusions of saline (n = 5), vehicle and low doses of Ro 60-0175 (27 & 90 μg kg-1 min1) evoked no significant changes in baseline EUS-EMG signal, urethral pressure and the micturition reflex in the case of the vehicles. However, Ro 60-0175 (27 μg kg-1 min-1) inhibited the micturition reflex, increasing volume (68 ± 10%) and pressure threshold (21 ± 8%), while at 90 & 270 μg kg-1 min-1 excitation was now observed. At 270 μg kg-1 min-1 Ro 60-0175 evoked EUS-EMG activity. Urethral and blood pressures and heart rate were unaffected. Infusion of Ro 60-0175 in the presence of CTE 4% had a similar effect on the above variables, although the high dose infusion caused a larger increase in the EUS-EMG signal, 1649 ± 178% c.f. 144 ± 11% and increased BP by 19 ± 1 mmHg. Similar effects were observed with Ro 60-0175 in the presence of SB 242084 (3.5, 22.5 & 67.5 μg kg-1 min-1); however the inhibitory action on micturition was reversed to excitation. In the presence of MDL (0.83 & 16.7 μg kg-1 min-1) Ro 60-0175 failed to evoke EUS EMG and attenuated the micturition reflex at all doses. A fall in BP was also observed. Infusion of the antagonists alone had no effect on the micturition variables, although the low dose of MDL caused a small attenuation (P<0.05) of the reflex. All doses of MDL caused a fall in BP.

These data indicate that activation of spinal 2A, not 2C receptors evokes EUS-EMG activity.

Y.M. was supported by a BBSRC PhD collaborative award with Pfizer UK. We also wish to thank Simon Westbrook, Gordon McMurray and Jennifer Gardiner at Pfizer UK for their support.

Mbaki Y and Ramage AG (2008) Br. J. Pharmacol 155, 343–356