Experimental Study On The Role Of Interstitial Cells Of Cajal(ICC) In Bladder Pacemaking

Spontaneous action potential can be recorded from muscular tissue of bladder, even if its nerval control is removed. That indicates that in some circumstance, spontaneous excitation and contraction of bladder could be induced by some'pacemaker cells'which remained unknown to us till now.Years ago, it was regarded that that'pacemaking'was from cells in detrusor but outcomes of a series of experiments showed that detrusor smooth muscle cells could not act as'pacemaker'due to the lack of their ability to evoke spontaneous excitation. Since that case, it was our goal to find which is the pacemaker of bladder and to indentify it.Study from gastrointestinal tract(GI) discovered that enterocinesia was induced by interstitial cells of Cajal(ICC) in it, so ICCs were regarded as pacemaker cells in GI. ICCs in GI, divided into different groups and reticulate, had quite a few structural junction not only to nerve terminal but also to smooth muscle cells.It was not long before the ICC-like cells was find in bladder, and the role of them was also thought to be similar to their counterpart in GI.From all accounts, study on ICC in bladder as pacemaker cell is still in primary phase. Evidences should be found to prove the hypothesis.The aim of the study is to investigate whether or not ICC in bladder is the pacemaker cell and we are going to pursue our goal from three directions: First, ICC as pacemaker should have a specilized structure. Characteristic of its distribution and structural connection between ICC and detrusor smooth muscle cell will be analysised in morphology.Second, ICC as pacemaker should have specilized function, that is,ICC could generate pacemaking current and transmit the current to neighboring detrusor smooth muscle cells. Third, detrusor function could be down-regulated when function of ICC was inhibited by blocking agent Glivec. Changes of detrusor function would be detected on the level of cell, tissue, and in vivo after blockade. Moreover, we made a intitial study on the structural and functional changes of ICC in instable bladder when the demonstration of ICC as pacemaker was accomplished.Results and conclusions:1. We established the effective method to investigate the morphology of ICC in the bladder: ICCs were identified to exist in suburothlium layer and the border of detrusor smooth muscle in bladder through KIT immunohistochemistry on the reformed stretch prepared bladder.2. We found the characteristic of ICC's distribution: ICCs exist as a network just like ICC in GI, which we found by immunofluorescence(IF).3. There was tight relationship between ICC and detrusor smooth muscle cell: ICCs were close to smooth muscle cells and they were intercrossed to each other when double-labeled IF was applied for analysis.Moreover,Gap junction was detected by transmission electron microscope(TEM).4. Hyperpolarization-activated cyclic nucleotide gate(HCN) ion channel, which was specified in generating pacemaking current, was found on the membrane of ICC: we used double-labeled IF by c-Kit and HCN antigen to detect it, so the capcity of ICC to generate pacemaking current was found.5. Signal transmission was found from ICC to detrusor smooth muscle cell: We found the cellar signal could be transmitted efficiently from ICC to detrusor smooth muscle cells using Fluorescence Recovery After Photobleaching (FRAP).That might be the functional basis for ICC to be pacemaker cell to initiate the action potential of detrusor smooth muscle cells.6. A mode for ICC's functional inhibition was established: The effects of blocking agent Glivec were mainly on the calcium influx of ICC which provided us a new mode to investigate the function of smooth muscle cells with ICC's function decreased.7. Changes of detrusor function were detected on the level of cell, tissue, and in in vivo. Calcium influx and signal transmission from ICC to detrusor were inhibited significantly and contractile amplitude of detrusor was also decreased significantly in muscle strip and bladder as a whole. That indicated that ICC would play a crucial important role in the movements of bladder. 8. Changes on ICC network and expression of HCN were found in instable detruor: Network in ICC was strengthened while the protein expression of HCN was increased significantly, which implied that changes of ICC may be the cause for functional abnormality of detrusor.