Biomechanical properties and innervation of the female caveolin‐1‐deficient detrusor

Abstract

BACKGROUND AND PURPOSECaveolin-1-deficiency is associated with substantial urogenital alterations. Here, a mechanical, histological and biochemical characterization of female detrusors from wild-type and caveolin-1-deficient (KO) mice was made to increase the understanding of detrusor changes caused by lack of caveolae. EXPERIMENTAL APPROACHLength-tension relationships were generated, and we recorded responses to electrical field stimulation, the muscarinic receptor agonist carbachol and the purinoceptor agonist ATP. Tyrosine nitration and the contents of caveolin-1, cavin-1, muscarinic M3 receptors, phospholipase Cb1, muscle-specific kinase (MuSK) and L-type Ca 2+ channels were determined by immunoblotting. Innervation was assessed by immunohistochemistry. KEY RESULTSBladder to body weight ratio was not changed, nor was there any change in the optimum circumference for force development. Depolarization-and ATP-induced stress was reduced, as was carbachol-induced stress between 0.1 and 3 mM, but the supramaximal relative (% K + ) response to carbachol was increased, as was M3 expression. The scopolamine-sensitive component of the electrical field stimulation response was impaired, and yet bladder nerves contained little caveolin-1. The density of cholinergic nerves was unchanged, whereas CART-and CGRP-positive nerves were reduced. Immunoblotting revealed loss of MuSK. CONCLUSIONS AND IMPLICATIONSAblation of caveolae in the female detrusor leads to generalized impairment of contractility, ruling out prostate hypertrophy as a contributing factor. Cholinergic neuroeffector transmission is impaired without conspicuous changes in the density of cholinergic nerves or morphology of their terminals, but correlating with reduced expression of MuSK. AbbreviationsCART, cocaine-and amphetamine-regulated transcript; Cav-1 or Cav1, caveolin-1; CaV1.2, pore-forming subunit of L-type Ca 2+ channel; CGRP, calcitonin gene-related peptide; EFS, electrical field stimulation; HK, K + -high solution (60 mM); HTX, Mayer's haematoxylin; KO, caveolin-1 knockout; L0, optimum circumference for force development; M3, type 3 muscarinic acetylcholine receptor; MuSK, muscle-specific kinase; NPK, neuropeptide K; PLCb1, phospholipase Cb1; VAChT, vesicular acetylcholine transporter; WT, wild type

Impaired contractility and detrusor hypertrophy in cavin-1-deficient mice

“…In detrusor myocytes they are arranged in parallel longitudinal membrane stripes and they reach a density of close to 2 caveolae per µm membrane (Murakumo et al, 1993;Sadegh et al, 2011). All caveolin isoforms are represented here, and it has been speculated that the different caveolins may play unique roles.…”

“…The length of the equatorial strip and the individual preparations was measured, preparations were cut, and loops were tied using the attached silk sutures. Length-tension relationships were obtained as described (Sadegh et al, 2011). Concentration-response curves were generated at the optimal circumference for force development (L 0 ) by cumulative addition of carbachol.…”

“…The animals were perfusion-fixed with 2.5 % glutaraldehyde in 150 mM cacodylate buffer (pH 7.4) through a carotid cannula. The bladders were dissected and transferred to new fixative, and then to cacodylate buffer and further prepared as described (Sadegh et al, 2011).…”

“…In some experiments detrusor strips from WT and cavin-1 KO mice were treated with or without 1 µM carbachol for 5 min and snap frozen in liquid nitrogen prior to protein extraction. Protein preparations were generated as described (Sadegh et al, 2011). 20 µg of protein was loaded on TGX Criterion gels (BioRad).…”

“…Work from several different laboratories including ours has demonstrated urogenital changes in caveolin-1 knock-out mice. These include impaired detrusor contractility and cholinergic neuroeffector transmission, bladder and prostate hypertrophy, enlargement of the seminal vesicles and disturbed erectile mechanisms (Lai et al, 2007;Lai et al, 2004;Sadegh et al, 2011;Shakirova et al, 2009;Woodman et al, 2004). Here we have used cavin-1 knock-out mice to explore the consequences of cavin-1 loss for detrusor structure, function and protein expression.…”