Urinary bladder function and somatic sensitivity in vasoactive intestinal polypeptide (VIP)-/- mice.

Vasoactive intestinal polypeptide (VIP) is an immunomodulatory neuropeptide widely distributed in neural pathways that regulate micturition. VIP is also an endogenous anti-inflammatory agent that has been suggested for the development of therapies for inflammatory disorders. In the present study, we examined urinary bladder function and hindpaw and pelvic sensitivity in VIP(-/-) and littermate wildtype (WT) controls. We demonstrated increased bladder mass and fewer but larger urine spots on filter paper in VIP(-/-) mice. Using cystometry in conscious, unrestrained mice, VIP(-/-) mice exhibited increased void volumes and shorter intercontraction intervals with continuous intravesical infusion of saline. No differences in transepithelial resistance or water permeability were demonstrated between VIP(-/-) and WT mice; however, an increase in urea permeability was demonstrated in VIP(-/-) mice. With the induction of bladder inflammation by acute administration of cyclophosphamide, an exaggerated or prolonged bladder hyperreflexia and hindpaw and pelvic sensitivity were demonstrated in VIP(-/-) mice. The changes in bladder hyperreflexia and somatic sensitivity in VIP(-/-) mice may reflect increased expression of neurotrophins and/or proinflammatory cytokines in the urinary bladder. Thus, these changes may further regulate the neural control of micturition.

Apical epidermal growth factor receptor signaling: regulation of stretch-dependent exocytosis in bladder umbrella cells.

The apical surface of polarized epithelial cells receives input from mediators, growth factors, and mechanical stimuli. How these stimuli are coordinated to regulate complex cellular functions such as polarized membrane traffic is not understood. We analyzed the requirement for growth factor signaling and mechanical stimuli in umbrella cells, which line the mucosal surface of the bladder and dynamically insert and remove apical membrane in response to stretch. We observed that stretch-stimulated exocytosis required apical epidermal growth factor (EGF) receptor activation and that activation occurred in an autocrine manner downstream of heparin-binding EGF-like growth factor precursor cleavage. Long-term changes in apical exocytosis depended on protein synthesis, which occurred upon EGF receptor-dependent activation of mitogen-activated protein kinase signaling. Our results indicate a novel physiological role for the EGF receptor that couples upstream mechanical stimuli to downstream apical EGF receptor activation that may regulate apical surface area changes during bladder filling.

ATP and purinergic receptor-dependent membrane traffic in bladder umbrella cells.

The umbrella cells that line the bladder are mechanosensitive, and bladder filling increases the apical surface area of these cells; however, the upstream signals that regulate this process are unknown. Increased pressure stimulated ATP release from the isolated uroepithelium of rabbit bladders, which was blocked by inhibitors of vesicular transport, connexin hemichannels, ABC protein family members, and nucleoside transporters. Pressure-induced increases in membrane capacitance (a measure of apical plasma membrane surface area where 1 microF approximately equals 1 cm2) were inhibited by the serosal, but not mucosal, addition of apyrase or the purinergic receptor antagonist PPADS. Upon addition of purinergic receptor agonists, increased capacitance was observed even in the absence of pressure. Moreover, knockout mice lacking expression of P2X2 and/or P2X3 receptors failed to show increases in apical surface area when exposed to hydrostatic pressure. Treatments that prevented release of Ca2+ from intracellular stores or activation of PKA blocked ATPgammaS-stimulated changes in capacitance. These results indicate that increased hydrostatic pressure stimulates release of ATP from the uroepithelium and that upon binding to P2X and possibly P2Y receptors on the umbrella cell, downstream Ca2+ and PKA second messenger cascades may act to stimulate membrane insertion at the apical pole of these cells.

Estrous cycle influences on sexual diergism of HPA axis responses to cholinergic stimulation in rats.

Central cholinergic systems differentially modulate hypothalamic-pituitary-adrenal (HPA) axis activity in female and male animals (sexual diergism). We previously reported that male rats had significantly greater HPA axis responses to stimulation by physostigmine (PHYSO), an acetylcholinesterase (AChE) inhibitor, compared to females. Females in defined estrous cycle stages, however, were not studied because of sample size limitations. We, therefore, determined HPA axis responses to stimulation by PHYSO in females during all estrous cycle stages (n = 78), and in male rats (n = 75). Plasma arginine vasopressin (AVP), adrenocorticotropic hormone (ACTH), and corticosterone (CORT) were measured. Estrous cycle stage was determined by light microscopy of vaginal smears. Proestrous and estrous females had higher ACTH and CORT responses compared to metestrous and diestrous females. Males had higher ACTH and AVP responses compared to females in all cycle stages. CORT responses followed the ACTH responses, except that females started from a higher baseline in all estrous stages, compared to males. These results suggest that cholinergic regulation of the HPA axis differs among females across stages of the estrous cycle, as well as between males and females. These effects are likely due to differences in circulating sex steroids and their actions within the brain.