Lipopolysaccharide stimulates BK channel activity in bladder umbrella cells

Abstract

Bladder urothelium plays an active role in response to bacterial infection. There is little known about the electrophysiological activity in urothelial cells in this process. We used a nonenzymatic method to isolate bladder urothelial tissue and to patch clamp umbrella cells in situ. A 200 pS conductance potassium (K) channel was detected from female C57BL6 mice. Of 58 total patches, 17.2% patches displayed the 200 pS K conductance channel. This K conductance channel showed Ca sensitivity and voltage dependence. Specific big-conductance potassium channel (BK) inhibitors (paxilline, iberiotoxin) blocked the 200 pS K conductance channel activity. RT-PCR and immunoblot confirmed BK channel pore-forming α-subunit (BK-α) mRNA and protein in urothelium. Immunohistochemistry also showed the BK-α located in urothelium. The above data provided evidence that the 200 pS K conductance channel was a BK channel. Lipopolysaccharide (LPS), a component of uropathogenic Escherichia coli, was used to investigate the role of BK channel in the pathogenesis of urinary tract infection. BK channel activity as NP increased threefold within 30 min of exposure to LPS. mRNAs for LPS receptors (TLR4, CD14, MD-2) were expressed in the urothelium but not in lamina propria or detrusor. Blockade of the receptors by an antagonist (polymyxin B) abrogated LPS's effect on BK channel. The involvement of protein kinase A (PKA) on BK channel activity was demonstrated by applying PKA blockers (H89 and PKI). Both PKA inhibitors abolished the BK channel activity induced by LPS. In conclusion, BK channel was identified in bladder umbrella cells, and its activity was significantly increased by LPS.

Sex differences in lower urinary tract biology and physiology

“…The translational relevance of these findings is that ERβ, and not ERα, is the estrogen receptor found in the bladder urothelium from female human subjects [ 26 ], and therefore, urothelial ERβ may play a more important role than ERα in UTI pathogenesis in women. A recent publication detected the presence of calcium-activated, voltage-gated, large conductance potassium channel (BK channel) on the umbrella urothelial cells [ 27 ]. Furthermore, urothelial BK channels were opened (activated) by lipopolysaccharides (LPS) suggesting that BK may work the same way in the urothelial cell as in macrophages where BK was important in regulating cytokine release when macrophages were exposed to LPS [ 28 ].…”

Mouse urothelial genes associated with voiding behavior changes after ovariectomy and bladder lipopolysaccharide exposure

“…Nine animals underwent sham and nine underwent OVX surgery. Four weeks after surgery, bladder urothelial tissues devoid of lamina propria were harvested ( n = 3 sham, n = 3 OVX mice) using the published technique . This was the “post‐surgery” or day 0 time point.…”

“…Four weeks after surgery, bladder urothelial tissues devoid of lamina propria were harvested (n = 3 sham, n = 3 OVX mice) using the published technique. 10 This was the "post-surgery" or day 0 time point. Intravesical LPS was instilled using the same protocol as shown in Figure 1 and urothelial tissues were harvested after LPS treatment on day 1 (n = 3 sham, n = 3 OVX mice) and on day 3 (n = 3 sham, n = 3 OVX mice).…”

Bladder urothelial BK channel activity is a critical mediator for innate immune response in urinary tract infection pathogenesis

“…Protocols PBS, LPS, and IBTX ϩ LPS were performed on another set of n ϭ 12 mice (4 for each protocol). After urine was collected at 2 h, the animals were euthanized, and a sheet of pure bladder urothelial tissue, free of lamina propria and detrusor, was harvested following the published protocol (14) and processed for qPCR experiments. These three inflammatory biomarkers were chosen because all three were significantly elevated after both LPS and UPEC exposure, and, furthermore, the elevations of these three cytokines/chemokines were also significantly blocked by IBTX for both LPS-and UPEC-exposed mice.…”

“…A recent publication demonstrated that, not only was calcium-activated voltage-gated potassium channel (BK channel) detected in the mouse umbrella cells, but BK channel activity (probability of open BK channel) was significantly increased by LPS exposure (14), suggesting that urothelial BK has a potential role in UTI pathogenesis. The role of BK in innate immune response has been shown in macrophages.…”