Adiponectin Signaling Regulates Urinary Bladder Function by Blunting Smooth Muscle Purinergic Contractility.

Unlabelled: Lower urinary tract symptoms (LUTS) affect ∼ 50% of the population aged >40 years and are strongly associated with obesity and metabolic syndrome. Adipose tissue plays a key role in obesity/metabolic syndrome by releasing adipokines that regulate systemic energy/lipid metabolism, insulin resistance, and inflammation. Adiponectin (ADPN), the most abundant adipokine, modulates energy/metabolism homeostasis through its insulin-sensitizing and anti-inflammatory effects.

DREADD agonist compound 21 causes acute diuresis in wild-type mice.

The targeted activation or inhibition of specific cell populations using chemogenetics allows the precise dissection of cellular signaling and function. Designer receptors exclusively activated by designer drugs (DREADDs) is a chemogenetic platform initially developed by mutating human muscarinic receptors to be unresponsive to endogenous acetylcholine but exclusively activated by an "inert" designer drug. Compound 21 (C21) is a new and potent DREADD agonist; however, radioligand assays from a recent report indicated its ability to bind to endogenous G protein-coupled receptors (GPCRs), including muscarinic M1-3 receptors.

Role of ecto-5'-nucleotidase in bladder function.

Purinergic signaling plays an important role in regulating bladder contractility and voiding. Abnormal purinergic signaling is associated with lower urinary tract symptoms (LUTS). Ecto-5'-nucleotidase (NT5E) catalyzes dephosphorylation of extracellular AMP to adenosine, which in turn promotes adenosine-A2b receptor signaling to relax bladder smooth muscle (BSM).

Minimizing the variables of voiding spot assay for comparison between laboratories.

The voiding spot assay (VSA) is increasingly being adopted as a standard method for assessing mouse urinary function. However, VSA outcomes are highly sensitive to housing environment and procedural parameters. Many variables exist among laboratories, including analytical software, type of daily housing cage, transportation, and the time of the day.

Deletion of Mechanosensory β1-integrin From Bladder Smooth Muscle Results in Voiding Dysfunction and Tissue Remodeling.

The bladder undergoes large shape changes as it fills and empties and experiences complex mechanical forces. These forces become abnormal in diseases of the lower urinary tract such as overactive bladder, neurogenic bladder, and urinary retention. As the primary mechanosensors linking the actin cytoskeleton to the extracellular matrix (ECM), integrins are likely to play vital roles in maintaining bladder smooth muscle (BSM) homeostasis.

Smooth Muscle Insulin Receptor Deletion Causes Voiding Dysfunction: A Mechanism for Diabetic Bladder Dysfunction.

Diabetic bladder dysfunction (DBD) is the most common complication in diabetes. Myogenic abnormalities are common in DBD; however, the underlying mechanisms leading to these remain unclear. To understand the importance of smooth muscle insulin receptor (IR)-mediated signaling in the pathogenesis of DBD, we conditionally deleted it to achieve either heterozygous (SMIR+/-) or homozygous (SMIR-/-) deletion in smooth muscle cells.

Reviving Cav1.2 as an attractive drug target to treat bladder dysfunction.

Inhibition of bladder contraction with antimuscarinics is a common approach to treat bladder hyperactivity, and the L-type voltage-gated calcium channel α (Cav1.2) is crucial for bladder contractility. Therefore, strategies aimed at inhibiting Cav1.

Molecular mechanisms of voiding dysfunction in a novel mouse model of acute urinary retention.

Acute urinary retention (AUR) is a common urological emergency and affects a significant patient population. The inability to eliminate urine may lead to permanent damage to the bladder's structure and functioning. However, we know little about the underlying molecular sequelae to the urine retention.

Disruption of Cav1.2-mediated signaling is a pathway for ketamine-induced pathology.

The general anesthetic ketamine has been repurposed by physicians as an anti-depressant and by the public as a recreational drug. However, ketamine use can cause extensive pathological changes, including ketamine cystitis. The mechanisms of ketamine's anti-depressant and adverse effects remain poorly understood.

CAKUT and Autonomic Dysfunction Caused by Acetylcholine Receptor Mutations.

Congenital anomalies of the kidney and urinary tract (CAKUT) are the most common cause of chronic kidney disease in the first three decades of life, and in utero obstruction to urine flow is a frequent cause of secondary upper urinary tract malformations. Here, using whole-exome sequencing, we identified three different biallelic mutations in CHRNA3, which encodes the α3 subunit of the nicotinic acetylcholine receptor, in five affected individuals from three unrelated families with functional lower urinary tract obstruction and secondary CAKUT. Four individuals from two families have additional dysautonomic features, including impaired pupillary light reflexes.

Targetable purinergic receptors P2Y12 and A2b antagonistically regulate bladder function.

Abnormalities in purine availability or purinergic receptor density are commonly seen in patients with lower urinary tract symptoms (LUTS), but the underlying mechanisms relating altered receptor function to LUTS are unknown. Here we provide extensive evidence for the reciprocal interplay of multiple receptors responding to ATP, ADP (adenosine diphosphate), and adenosine, agonists that regulate bladder function significantly. ADP stimulated P2Y12 receptors, causing bladder smooth muscle (BSM) contraction, whereas adenosine signaling through potentially newly defined A2b receptors, actively inhibited BSM purinergic contractility.

Role of P2X Receptor in Mouse Voiding Function.

Purinergic signalling plays an important role in the regulation of bladder smooth muscle (BSM) contractility, and P2X receptor is expressed in the bladder wall, where it may act by forming heteromeric receptors with P2X, the major purinergic force-generating muscle receptor. To test this hypothesis, we examined mouse BSM contractile properties in the absence and presence of selective P2X (NF449 & NF279) and P2X antagonists (5-BDBD). These drugs inhibited BSM purinergic contraction only partially, suggesting the possibility of a heteromeric receptor.

Evaluating the voiding spot assay in mice: a simple method with complex environmental interactions.

The voiding spot assay (VSA) on filter paper is an increasingly popular method for studying lower urinary tract physiology in mice. However, the ways VSAs are performed differ significantly between laboratories, and many variables are introduced compared with the mouse's normal housing situation. Rodents are intelligent social animals, and it is increasingly understood that social and environmental stresses have significant effects on their physiology.

Intact urothelial barrier function in a mouse model of ketamine-induced voiding dysfunction.

Ketamine is a popular choice for young drug abusers. Ketamine abuse causes lower urinary tract symptoms, with the underlying pathophysiology poorly understood. Disruption of urothelial barrier function has been hypothesized to be a major mechanism for ketamine cystitis, yet the direct evidence of impaired urothelial barrier function is still lacking.

Polarized ATP distribution in urothelial mucosal and serosal space is differentially regulated by stretch and ectonucleotidases.

Purinergic signaling is a major pathway in regulating bladder function, and mechanical force stimulates urothelial ATP release, which plays an important role in bladder mechanotransduction. Although urothelial ATP release was first reported almost 20 years ago, the way in which release is regulated by mechanical force, and the presence of ATP-converting enzymes in regulating the availability of released ATP is still not well understood. Using a set of custom-designed Ussing chambers with the ability to manipulate mechanical forces applied on the urothelial tissue, we have demonstrated that it is stretch and not hydrostatic pressure that induces urothelial ATP release.

Evaluation of voiding assays in mice: impact of genetic strains and sex.

Void spot assays (VSA) and cystometry are two of the most common tests performed in mice to assess lower urinary tract function. Assay protocols and methodology vary greatly among laboratories, and little is known about reproducibility of results generated by different laboratories. We performed VSA in four mouse strains, comparing males with females and comparing results between two independent laboratories.

Hypercompliant apical membranes of bladder umbrella cells.

Urinary bladder undergoes dramatic volume changes during filling and voiding cycles. In the bladder the luminal surface of terminally differentiated urothelial umbrella cells is almost completely covered by plaques. These plaques (500 to 1000 nm) are made of a family of proteins called uroplakins that are known to form a tight barrier to prevent leakage of water and solutes.

ADP-induced bladder contractility is mediated by P2Y12 receptor and temporally regulated by ectonucleotidases and adenosine signaling.

Purinergic signaling comprises one key pathway in modulating bladder smooth muscle (BSM) contractility, disorders of which become highly prevalent with aging. ADP was first observed to modulate BSM contractility >40 yr ago, yet the underlying molecular mechanism still remains unclear. Here, we demonstrate, using myography, that ADP and ADPβS dose-dependently induce mouse BSM contraction, and ADP-induced BSM contraction is blocked by a selective P2Y12 receptor (P2Y12R) antagonist, PSB 0739 (25 μM), but is unaffected by P2Y1 and P2Y13 receptor antagonists.

Spontaneous voiding by mice reveals strain-specific lower urinary tract function to be a quantitative genetic trait.

Lower urinary tract (LUT) symptoms become prevalent with aging and affect millions; however, therapy is often ineffective because the etiology is unknown. Existing assays of LUT function in animal models are often invasive; however, a noninvasive assay is required to study symptom progression and determine genetic correlates. Here, we present a spontaneous voiding assay that is simple, reproducible, quantitative, and noninvasive.

Lack of specificity shown by P2Y6 receptor antibodies.

P2Y6 receptor in bladder smooth muscle responds to UDP by increasing muscle tone and augmenting bladder contractions. The exact cellular location of the receptor is however unknown. Three commercially available antibodies to P2Y6 receptor gave clean bands on Western blot which were eliminated by specific peptide competition.

Loss of β1-integrin from urothelium results in overactive bladder and incontinence in mice: a mechanosensory rather than structural phenotype.

Bladder urothelium senses and communicates information about bladder fullness. However, the mechanoreceptors that respond to tissue stretch are poorly defined. Integrins are mechanotransducers in other tissues.

Extracellular UDP enhances P2X-mediated bladder smooth muscle contractility via P2Y(6) activation of the phospholipase C/inositol trisphosphate pathway.

Bladder dysfunction characterized by abnormal bladder smooth muscle (BSM) contractions is pivotal to the disease process in overactive bladder, urge incontinence, and spinal cord injury. Purinergic signaling comprises one key pathway in modulating BSM contractility, but molecular mechanisms remain unclear. Here we demonstrate, using myography, that activation of P2Y6 by either UDP or a specific agonist (MRS 2693) induced a sustained increase in BSM tone (up to 2 mN) in a concentration-dependent manner.

Cellular expression profile for interstitial cells of cajal in bladder - a cell often misidentified as myocyte or myofibroblast.

Background: Interstitial cells of Cajal (ICC) have been identified in urinary bladder of several species, but their presence in mice remains uncertain. Meanwhile, dozens of reports indicate that dysregulation of connexin 43 plays an important role in bladder overactivity, but its localization has not been clearly defined, with reports of expression in either the smooth muscle or in myofibroblasts. We recently identified a population of ectonucleoside triphosphate diphosphohydrolase 2 (NTPDase2) positive cells that resemble ICC and are distinct from smooth muscle, fibroblasts, myofibroblasts and neurons.