Circadian clocks are the endogenous oscillators that harmonize a variety of physiological processes within the body. Although many urinary functions exhibit clear daily or circadian variation in diurnal humans and nocturnal rodents, the precise mechanisms of these variations are as yet unclear. In the present study, we demonstrate that Per2 promoter activity clearly oscillates in neonate and adult bladders cultured ex vivo from Per2::Luc knock-in mice. In subsequent experiments, we show that multiple local oscillators are operating in all the bladder tissues (detrusor, sphincter and urothelim) and the lumbar spinal cord (L4-5) but not in the pontine micturition center or the ventrolateral periaqueductal gray of the brain. Accordingly, the water intake and urine volume exhibited daily and circadian variations in young adult wild-type mice but not in Per1-/- Per2-/- mice, suggesting a functional clock-dependent nature of the micturition rhythm. Particularly in PDK mice, the water intake and urinary excretion displayed an arrhythmic pattern under constant darkness, and the amount of water consumed and excreted significantly increased compared with those of WT mice. These results suggest that local circadian clocks reside in three types of bladder tissue and the lumbar spinal cord and may have important roles in the circadian control of micturition function.
Animals ; *Circadian Clocks ; Drinking ; Mice ; Organ Specificity ; Periaqueductal Gray/metabolism/physiology ; Period Circadian Proteins/genetics/*metabolism ; Pons/metabolism/physiology ; Spinal Cord/*metabolism/physiology ; Urinary Bladder/innervation/metabolism/*physiology ; Urination

The antisense approach and RT-PCR were used to study the effects of muscarinic receptors on the scores of morphine-withdrawal syndrome and the expression of NMDA receptor subtypes (NR(1A) and NR(2A)) mRNA in rat spinal cord and brainstem. The concentrations of glutamate in periaqueductal grey (PAG) of morphine-withdrawal rats were determined by capillary electrophoresis with laser-induced fluorescence detection. The data showed that the NR(1A) and NR(2A) mRNA levels were increased significantly in the spinal cord and brainstem 1 h after the injection of naloxone (4 mg/kg, i.p.) in morphine-dependent rats. Moreover, in morphine-dependent rats pretreated (i.p.) with scopolamine (0.5 mg/kg), or pirenzepine (10 mg/kg), MK801 (0.125 mg/kg), L-N-nitroarginine methylester (10 mg/kg) 30 min before naloxone injection, the NR(1A) and NR(2A) mRNA levels were significantly lower than those of 1 h morphine-withdrawal rats. Intrathecal injection of NR(1A) or M(2) receptor antisense oligonucleotides (A-oligo, 4 microg/per rat) 24 h prior to naloxone challenge could block the morphine withdrawal symptoms including wet dog shaking, irritability, salivation, diarrhea, chewing and weight loss. Meanwhile, in morphine-dependent rats the NR(1A) mRNA levels in the spinal cord and brainstem were down-regulated by intrathecal injection of M(2) receptor A-oligo. The glutamate concentrations in PAG microdialysis were increased to a maximal level 15 min after naloxone injection. The glutamate response was inhibited by pretreatment with M(2) receptor A-oligo but not by M(1) A-oligo. The results suggest that the expression of NMDA receptors and the release of glutamate in brainstem are involved in the processes of morphine withdrawal and that the NMDA receptor expression is possibly regulated by the muscarinic receptors during morphine withdrawal.
Animals ; Brain Stem ; metabolism ; Glutamic Acid ; metabolism ; Male ; Morphine ; adverse effects ; Periaqueductal Gray ; metabolism ; physiology ; Rats ; Rats, Sprague-Dawley ; Receptors, Muscarinic ; physiology ; Receptors, N-Methyl-D-Aspartate ; biosynthesis ; genetics ; Spinal Cord ; metabolism ; Substance Withdrawal Syndrome ; genetics ; metabolism

OBJECTIVETo investigate the role of ventrolateral periaqueductal gray (VL-PAG) metabotropic glutamate receptors subtype 7 and 8 (mGluR 7/8) in descending modulation of cardiosomatic motor reflex (CMR) in rats.

METHODSAMN082 (agonist of mGluR 7) and DCPG (agonist of mGluR 8) were injected into the VL-PAG of a rat model of CMR to observe their effects in modulating CMR. The raphe magnus nucleus (NRM) or the gigantocellular reticular nucleus (Gi) was then damaged, and the changes in VL-PAG descending modulation were observed.

RESULTSSelective activation of mGluR 7 of the VL-PAG by AMN082 obviously facilitated capsaicin (CAP)-induced CMR (P<0.05), which was suppressed by DCPG-induced mGluR 8 activation (P<0.05). These facilitatory or inhibitory effects were completely reversed by group III mGluR antagonist MSOP. Damaging the NRM of VL-PAG main relay nucleus did not significantly affect the facilitatory effect produced by AMN082 microinjection (P>0.05), but partially attenuated the inhibitory effect of DCPG microinjection (P<0.05). Both the facilitatory effect of AMN082 and the inhibitory effect of DCPG were reduced obviously after bilateral Gi damage (P<0.05).

CONCLUSIONVL-PAG mGluR 7 and mGluR 8 mediate biphasic regulation of CMR in rats probably through activation of different sub-nuclei and different neurons in the rostroventral medulla.

Animals ; Benzhydryl Compounds ; pharmacology ; Benzoates ; pharmacology ; Glycine ; analogs & derivatives ; pharmacology ; Male ; Medulla Oblongata ; metabolism ; Periaqueductal Gray ; metabolism ; Physical Conditioning, Animal ; Rats ; Rats, Sprague-Dawley ; Receptors, Metabotropic Glutamate ; agonists ; metabolism ; Reflex ; physiology