AB287. SPR-14 High fat diet leads to impaired mitochondrial respiration, increased hydrogen peroxide emission and greater nerve-mediated purinergic contraction in mouse bladder

Objective: Chronic high fat diet (HFD) is associated with the development of metabolic syndrome and type 2 diabetes, known risk factors for lower urinary tract symptoms, overactive bladder, and incontinence. HFD induced increases in oxidative stress and impaired mitochondrial integrity is a possible mechanism underlying this pathology. Our objectives were to determine bladder function following long-term HFD and characterize mitochondrial oxidant emission and respiratory function. We hypothesized that chronic HFD would lead to increased cholinergic mediated contraction and impaired mitochondrial respiration.

Methods: Male C57BL/6J mice were maintained on a control diet (CD: 4% fat, 60% carbohydrates, 26% protein) or a HFD (45% fat, 35% carbohydrates, 20% protein) from 6 weeks of age for 20 weeks. A week prior to sacrifice, lower urinary tract function was assessed via void spot assay. Animals were euthanized and bladders and visceral fat depots were collected and weighed. Bladders were denuded of urothelium, cut into 8×2 mm strips and placed in tissue baths. Increasing concentrations of carbachol (cholinergic mediated) contraction and adrenergic relaxation to norepinephrine (NE) were measured. Electric field stimulated (EFS) contractions with and without atropine (cholinergic antagonist) were assessed. A separate group of bladders were denuded, homogenized and mitochondria quantified via citrate synthase activity. High-resolution respirometry measured oxygen consumption of individual electron transport chain complexes. Mitochondrial hydrogen peroxide (H₂O₂) emission was measured fluorimetrically using Amplex Ultra Red. Data were compared using a student t-test.

Results: HFD mice had a greater number of voids (P<0.005) and a larger void area (P<0.005). Blood glucose was greater in HFD (CD: 85±6.1; HFD: 139±5.9) as were body and overall visceral fat weights (P<0.0005). In contrast, HFD did not impact bladder weight. HFD did not change detrusor cholinergic contraction or adrenergic relaxation. However, detrusor contraction to EFS was markedly increased with HFD in both the presence and absence of atropine. Cholinergic inhibition blunted EFS contraction to a greater extent in CD bladders (CD: 75.7%±2.65; HFD: 42.3%±2.82, P<0.05) indicating that HFD had shifted EFS contraction to greater purinergic release. Citrate synthase activity indicated similar amounts of intact mitochondria. HFD resulted in a decreased respiratory capacity across 4 of 6 mitochondrial complexes investigated (P<0.05). Complex I and Complex II in state 4 conditions were unchanged. Additionally, mitochondrial H₂O₂ emission was increased in HFD bladders (CD: 11.4 pmols/min/mg ±0.53; HFD: 18.6±0.66, P<0.05).

Conclusions: Prolonged HFD leads to increased urinary frequency and volume, increased detrusor purinergic EFS contraction and mitochondrial dysfunction including decreased respiratory capacity and increased H₂O₂ emission. Further investigation of the role of mitochondrial dysfunction and resulting detrusor dysfunction following HFD is merited.

Funding Source(s): None

Keywords: Mitochondria; bladder; high fat diet (HFD); in vitro contractility; purinergic