Nerve growth factor (NGF) has been proposed as an important mediator inducing bladder overactivity under pathological conditions such as spinal cord injury, bladder outlet obstruction, or cystitis. We therefore examined the effects of chronic NGF treatment on bladder activity and the properties of bladder afferent neurons. In adult female rats, NGF (2.5 μg/μl) was infused continuously into the intrathecal space at the L6–S1 level of spinal cord for 1 or 2 weeks using osmotic pumps (0.5 μl/h). Bladder afferent neurons were labeled with axonal transport of Fast Blue injected into the bladder wall. After intrathecal injection of NGF, cystometrograms under an awake condition showed bladder overactivity revealed by time-dependent reductions in intercontraction intervals and voided volume. ELISA analyses showed significant increases in NGF levels in L6–S1 dorsal root ganglia of NGF-treated rats. In patch-clamp recordings, dissociated bladder afferent neurons exhibiting tetrodotoxin (TTX)-resistant action potentials from NGF-treated animals were larger in diameter and had significantly lower thresholds for spike activation compared with sham rats. In addition, the number of TTX-resistant action potentials during 600 ms depolarizing pulses was significantly increased time dependently after 1 or 2 weeks of NGF application. The density of slowly inactivating A-type K⁺currents was decreased by 52% in bladder afferent neurons with TTX-resistant spikes after 2 week NGF treatment. These results indicate that increased NGF levels in bladder afferent pathways and NGF-induced reduction in A-type K⁺current density could contribute to the emergence of bladder overactivity as well as somal hypertrophy and hyperexcitability of bladder afferent neurons.

The present study investigated the effect of inhibiting the expression of Nav1.8 (PN3/SNS) sodium channels by an antisense oligodeoxynucleotide (ODN) on bladder nociceptive responses induced by intravesical acetic acid infusion in rats. Animals were injected intrathecally with either Nav1.8 antisense or mismatch ODN. Control cystometrograms under urethane anesthesia during intravesical saline infusion exhibited intercontraction intervals (ICIs) that were significantly longer in antisense-treated rats than in mismatch ODN-treated rats. Intravesical infusion of 0.1% acetic acid induced bladder hyperactivity as reflected by a 68% reduction in ICIs in mismatch ODN-treated rats but did not significantly reduce ICIs in antisense-treated rats. The number of Fos-positive cells after acetic acid administration were significantly reduced in the L6 spinal cord from antisense-treated animals, compared with mismatch ODN-treated animals. In addition, Nav1.8 immunoreactivity was reduced in L6 dorsal root ganglion neurons in the antisense-treated rat. In patch-clamp recordings, the conductance density of TTX-resistant sodium currents in dissociated bladder afferent neurons that were labeled by axonal transport of a fluorescent dye, Fast Blue, injected into the bladder wall was also smaller in antisense-treated rats than in mismatch ODN-treated rats, whereas no changes were observed in TTX-sensitive currents. These results indicate that the Nav1.8 TTX-resistant sodium channels are involved in the activation of afferent nerves after chemical irritation of the bladder. These channels represent a new target for the treatment of inflammatory pain from visceral organs such as the urinary bladder.

Bladder pain syndrome/interstitial cystitis is a disease with lower urinary tract symptoms, such as bladder pain and urinary frequency, which results in seriously impaired quality of life of patients. The extreme pain and urinary frequency are often difficult to treat. Although the etiology of bladder pain syndrome/interstitial cystitis is still not known, there is increasing evidence showing that afferent hyperexcitability as a result of neurogenic bladder inflammation and urothelial dysfunction is important to the pathophysiological basis of symptom development. Further investigation of the pathophysiology will lead to the effective treatment of patients with bladder pain syndrome/interstitial cystitis.

The effect of a nitric oxide (NO) donor on high-voltage-activated Ca²⁺channel currents ( I_Ca) was examined using the whole cell patch-clamp technique in L₆–S₁dorsal root ganglion (DRG) neurons innervating the urinary bladder. The neurons were labeled by axonal transport of a fluorescent dye, Fast Blue, injected into the bladder wall. Approximately 70% of bladder afferent neurons exhibited tetrodotoxin (TTX)-resistant action potentials (APs), and 93% of these neurons were sensitive to capsaicin, while the remaining neurons had TTX-sensitive spikes and were insensitive to capsaicin. The peak current density of nimodipine-sensitive L-type Ca²⁺channels activated by depolarizing pulses (0 mV) from a holding potential of −60 mV was greater in bladder afferent neurons with TTX-resistant APs (39.2 pA/pF) than in bladder afferent neurons with TTX-sensitive APs (28.9 pA/pF), while the current density of ω-conotoxin GVIA-sensitive N-type Ca²⁺channels was similar (43–45 pA/pF) in both types of neurons. In both types of neurons, the NO donor, S-nitroso- N-acetylpenicillamine (SNAP) (500 μM), reversibly reduced (23.4–26.6%) the amplitude of I_Caelicited by depolarizing pulses to 0 mV from a holding potential of −60 mV. SNAP-induced inhibition of I_Cawas reduced by 90% in the presence of ω-conotoxin GVIA but was unaffected in the presence of nimodipine, indicating that NO-induced inhibition of I_Cais mainly confined to N-type Ca²⁺channels. Exposure of the neurons for 30 min to 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10 μM), an inhibitor of NO-stimulated guanylyl cyclase, prevented the SNAP-induced reduction in I_Ca. Extracellular application of 8-bromo-cGMP (1 mM) mimicked the effects of NO donors by reducing the peak amplitude of I_Ca(28.6% of reduction). Action potential configuration and firing frequency during depolarizing current pulses were not altered by the application of SNAP (500 μM) in bladder afferent neurons with TTX-resistant and -sensitive APs. These results indicate that NO acting via a cGMP signaling pathway can modulate N-type Ca²⁺channels in DRG neurons innervating the urinary bladder.