In the present study it was particularly
In the present study, it was particularly of interest that intravesical administration of PGE2 evoked ONO-8130-reversible rapid phosphorylation of ERK in the L6 spinal cord. Detection of phosphorylated ERK is now often used as a marker for somatic and visceral nociception, including p38 inhibitors pain , , . MDH, LDH, DCM, and SPN in the L6 spinal cord where phosphorylation of ERK occurred following intravesical PGE2 in the present study, correspond to the spinal regions that exhibit delayed expression of Fos protein after irritation of the lower urinary tract , . Those areas receive afferent projections from the urinary bladder , . EP1 receptors are known to be expressed in capsaicin-sensitive dorsal root ganglion neurons, and activation of the neuronal EP1 receptors causes sensitization of transient receptor potential vanilloid-1 (TRPV1) channels . On the other hand, EP1 receptors are also expressed in the urothelium, and activation of urothelial EP1 receptors facilitates adenosine triphosphate release from urothelial cells . The released adenosine triphosphate is capable of activating P2X3 receptors in the primary afferent terminals in the bladder to convey sensory information to the central nervous system . Together, it is likely that both neural and urothelial EP1 receptors mediate the PGE2-induced phosphorylation of ERK in the spinal cord. In the urinary bladder, other PGE2 receptors, EP2, EP3, and EP4, in addition to EP1, might modulate bladder functions in health and disease , , , . Our finding that ONO-8130 exerted only partial inhibition on ERK phosphorylation in MDH and SPN (Fig. 6) might suggest involvement of PGE2 receptors other than EP1. Our study to identify PGE2 receptor subtypes, particularly EP4, involved in the ONO-8130-resistant component of intravesical PGE2-evoked ERK phosphorylation, is now in progress. As reported previously , , , systemic administration of cyclophosphamide evoked upregulation of COX-2 at mRNA and protein levels in the bladder tissue in the present study, suggesting increased prostanoid formation in association with the development of cystitis. Nevertheless, diclofenac, an inhibitor of COX, was less effective against bladder pain in cyclophosphamide-treated mice, compared with ONO-8130 (Fig. 3, Fig. 4B). There is plenty of evidence that nonsteroidal antiinflammatory drugs produce damage to the bladder epithelium or cystitis , , , , implying the protective role of certain endogenous prostanoids in the urothelium. In this context, selective antagonism of EP1 receptors by ONO-8130 is considered profitable for treatment of cystitis-related pain, compared with total inhibition of production of both pronociceptive/proinflammatory and antiinflammatory prostanoids by nonsteroidal antiinflammatory drugs. Our findings that pretreatment with AH6809, an EP2 antagonist, had no effect on the cyclophosphamide-induced bladder pain and increased bladder weight, suggest that EP2 receptors are not involved in this cystitis model. Most recently, it has been reported that EP4 receptors were upregulated in the bladder and spinal cord, after cyclophosphamide was administered repeatedly, every 3days . Therefore, it is likely that EP4, in addition to EP1, might participate in the bladder pain/inflammation induced by cyclophosphamide, and our ongoing study focuses on the roles of EP4 receptors in the acute and chronic/sub-chronic phases of the cyclophosphamide-induced cystitis. The major symptoms in interstitial cystitis patients include increased voiding frequency in addition to bladder pain. The hyperactive voiding is considered to be associated with activation/sensitization of bladder afferents. Although the effect of the EP1 antagonist on the increased voiding reflex and/or hyperactivity of bladder afferents in the cyclophosphamide-induced cystitis model has yet to be examined, there is evidence that EP1 antagonists inhibit the neuronal activity after intravesical administration of acetic acid in rats  and reverse the shortened micturition interval in rats with bladder outlet obstruction . Further, in cystometrogram, EP1 agonists shorten the intercontraction interval in wild-type, but not EP1-knockout, mice . Thus, EP1 receptors appear to play critical roles in increased voiding activity as well as bladder pain, suggesting that EP1 receptor antagonists may reduce both bladder pain and increased voiding reflex in patients with interstitial cystitis.