MP60-06 PATHOGENIC ROLE OF TRUNCATED TRKB RECEPTOR ISOFORM (TRKB.T1) IN BDNF INDUCED DETRUSOR OVERACTIVITY (DO)

Abstract

INTRODUCTION AND OBJECTIVES: Overexpression of brain derived neurotrophic factor (BDNF) is well known in overactive bladder (OAB). Amelioration of cyclophosphamide induced DO by antibody mediated sequestration of BDNF highlighted the pathogenic role of BDNF receptors and their downstream signaling. Tropomyosin receptor kinase B (TrkB) is the high affinity receptor for BDNF, which is either expressed as a full length receptor (TrkB FL) or as a splice variant called truncated, TrkB (TrkB.T1). Here, we tested whether gene silencing targeted for TrkB.T1 can overcome the BDNF induced DO. METHODS: Under isoflurane anesthesia, the bladder wall of female Sprague-dawley rats was injected with BDNF plasmid complexed with liposomes (n1⁄45) alone (10mg) or together with either antisense (n1⁄45) or scrambled oligonucleotide against TrkB.T1 (n1⁄45). 7 days later, cystometry was performed followed by bladder harvest to analyze the expression of small Rho GTPase and phosphorylation of Myosin light chain (MLC). RESULTS: As reported earlier, bladder wall injection of BDNF plasmid complexed with liposomes causes BDNF overexpression and DO is observed 7 days later. BDNF induced DO is evident from reduced intercontractile intervals (ICI) and voided volume in rat group injected with BDNF plasmid + scrambled oligonucleotide. In contrast, gene silencing of TrkB.T1 by antisense significantly reversed the DO and increased the ICI and voided urine volume. DO in the scrambled group was associated with the higher expression of Rho kinases viz., RhoA, Cdc42, Rac1 along with the increase phosphorylation of pMLC at Th18 and Ser 19 position. Gene silencing of TrkB.T1 reduced the expression of Rho kinases and completely blocked the phosphorylation of MLC at both positions. CONCLUSIONS: Here, we report that BDNF induced DO is primarily driven by the dominant negative characteristic of TrkB.T1. The tyrosine kinase domain of full length TrkB FL is missing in TrkB.T1, but there exists a domain which interacts with RhoGDi proteins and regulates cell contractility via Rho kinase-mediated increase in phosphorylation of MLC. The C-terminal sequence of TrkB is completely conserved in mammals and therefore verification of TrkB.T1 expression in bladder biopsy of OAB patients is the appropriate next step to establish it as a drug target for treating voiding dysfunctions.