Urothelium muscarinic activation phosphorylates CBS(Ser227) via cGMP/PKG pathway causing human bladder relaxation through H(2)S production

The urothelium modulates detrusor activity through releasing factors whose nature has not been clearly defined. Here we have investigated the involvement of H(2)S as possible mediator released downstream following muscarinic (M) activation, by using human bladder and urothelial T24 cell line. Carbachol stimulation enhances H(2)S production and in turn cGMP in human urothelium or in T24 cells. This effect is reversed by cysthationine-β-synthase (CBS) inhibition. The blockade of M1 and M3 receptors reverses the increase in H(2)S production in human urothelium. In T24 cells, the blockade of M1 receptor significantly reduces carbachol-induced H(2)S production. In the functional studies, the urothelium removal from human bladder strips leads to an increase in carbachol-induced contraction that is mimicked by CBS inhibition. Instead, the CSE blockade does not significantly affect carbachol-induced contraction. The increase in H(2)S production and in turn of cGMP is driven by CBS-cGMP/PKG-dependent phosphorylation at Ser(227) following carbachol stimulation. The finding of the presence of this crosstalk between the cGMP/PKG and H(2)S pathway downstream to the M1/M3 receptor in the human urothelium further implies a key role for H(2)S in bladder physiopathology. Thus, the modulation of the H(2)S pathway can represent a feasible therapeutic target to develop drugs for bladder disorders.