Role for pituitary adenylate cyclase activating polypeptide in cystitis-induced plasticity of micturition reflexes

Abstract

Pituitary adenylate cyclase activating polypeptide (PACAP) peptides are expressed and regulated in sensory afferents of the micturition pathway. Although these studies have implicated PACAP in bladder control, the physiological significance of these observations has not been firmly established. To clarify these issues, the roles of PACAP and PACAP signaling in micturition and cystitis were examined in receptor characterization and physiological assays. PACAP receptors were identified in various tissues of the micturition pathway, including bladder detrusor smooth muscle and urothelium. Bladder smooth muscle expressed heterogeneously PAC 1null, PAC1HOP1, and VPAC2 receptors; the urothelium was more restricted in expressing preferentially the PAC 1 receptor subtype only. Immunocytochemical studies for PAC 1 receptors were consistent with these tissue distributions. Furthermore, the addition of 50 -100 nM PACAP27 or PACAP38 to isolated bladder strips elicited transient contractions and sustained increases in the amplitude of spontaneous phasic contractions. Treatment of the bladder strips with tetrodotoxin (1 M) did not alter the spontaneous phasic contractions suggesting direct PACAP effects on bladder smooth muscle. PACAP also increased the amplitude of nerve-evoked contractions. By contrast, vasoactive intestinal polypeptide had no direct effects on bladder smooth muscle. In a rat cyclophosphamide (CYP)-induced cystitis paradigm, intrathecal or intravesical administration of PAC 1 receptor antagonist, PACAP6 -38, reduced cystitis-induced bladder overactivity. In summary, these studies support roles for PACAP in micturition and suggest that inflammation-induced plasticity in PACAP expression in peripheral and central micturition pathways contribute to bladder dysfunction with cystitis.neuropeptides; urinary bladder; bladder overactivity; dorsal root ganglia; spinal cord; inflammation THE STORAGE AND PERIODIC ELIMINATION of urine requires a complex neural control system that coordinates the activities of the smooth muscle of the urinary bladder and the smooth and striated muscle of the urethral sphincters (13,32,33). Coordination between these organs is mediated by a complex neural control system located in the brain, spinal cord, and peripheral ganglia (12). Experiments with a chemically (cyclophosphamide, CYP)-induced bladder inflammation (11, 34,38) rodent model have demonstrated alterations in neurochemical (52, 53, 66, 69), electrophysiological (29, 72), organizational (65, 68), and functional properties of the micturition reflex (24, 38, 39), suggesting dramatic reorganization of the micturition reflex pathways. Alterations in peripheral bladder afferent (sensory)/ efferent (autonomic and motor) and central interneuronal pathway functions may underlie detrusor overactivity that accompany CYP-induced cystitis.Pituitary adenylate cyclase activating polypeptide (PACAP) peptides have diverse functions in the endocrine, nervous, gastrointestinal, and cardiovascular systems (1, 6) through PAC 1 , VPAC 1 and VPAC ...

Expression of corticotropin-releasing factor and CRF receptors in micturition pathways after cyclophosphamide-induced cystitis

“…We have confirmed that the CRF IR in the lateral horn of L 6 -S 1 is associated with the autonomic nucleus by using segmentation protocols previously used and validated (6,61,64) and by also staining spinal cord sections for nNOS IR (48), a marker for preganglionic autonomic neurons (Fig. 4C).…”

“…Previous studies have demonstrated that a number of neurotrophic factors (15,59), neuropeptides (6,57), and cytokines (27,67) are upregulated in the urinary bladder after CYPinduced cystitis. Subtractive techniques to block these factors improve bladder function and reduce pain-related behaviors (6, (A and B) or nerve fibers in the suburothelial plexus in whole mount preparations of the urothelium/suburothelium (C and D) in control (A and C) or CYP-treated (B and D) rats.…”

“…In this study, use of the word urothelium refers to the urothelium and suburothelial structures. Total RNA from the different tissues was extracted using the STAT-60 total RNA/mRNA isolation reagent (Tel-Test "B," Friendswood, TX), as previously described (6). The quality and quantity of the resulting RNA were assessed with spectrometry at 260/280 nm, and 2 g of total RNA from each sample were used to synthesize first-strand cDNA using SuperScript II RT with the SuperScript II Preamplification System (Invitrogen, Carlsbad, CA) in a 20-l final reaction volume.…”

“…These changes may be mediated by chemical mediators (e.g., neurotrophins, cytokines, neuropeptides) produced in the bladder, spinal cord, or dorsal root ganglia with cystitis (5,27,57,59,61).…”

Expression of Phosphorylated cAMP Response Element Binding Protein (p-CREB) in Bladder Afferent Pathways in VIP−/− Mice with Cyclophosphamide (CYP)-Induced Cystitis

“…Greater basal activity in the primary bladder afferents in the DRG or in the target organ itself (i.e., urinary bladder) could result in increased pCREB expression in the absence of inflammation by increasing Ca ++ and/or cAMP in VIP −/− mice. Primary afferent cells in the DRG express VIP (Morgan et al, 1999;Hernandez et al, 2006) and VIP receptors (Yashpal et al, 1991;Braas et al, 2006). Thus, absence of VIP in the VIP −/− mice could produce a direct effect on primary afferent cells.…”