Willmann Liang

Three months of methoxetamine administration is associated with significant bladder and renal toxicity in mice

Abstract Context. Methoxetamine is a ketamine analogue that has recently emerged as a novel psychoactive substance. Chronic ketamine use is associated with significant bladder and renal toxicity. Methoxetamine has been marketed as “bladder friendly”, but there is no data to be able to substantiate this claim. Objective. To characterise the patterns of bladder and renal toxicity associated with 3 months of methoxetamine administration in an animal model. Materials and methods. Two-month-old Institute of Cancer Research mice were administered 30 mg/kg methoxetamine intraperitoneally (n = 5) or saline (n = 3 control) for 3 months. The animals were then sacrificed and histological examination, immuno-cytochemistry using polyclonal anti-CD4 antibodies and sirius-red staining for collagen were performed. Results. The kidneys of methoxetamine-treated animals showed inflammatory cell infiltration, tubular cell necrosis and glomerular damage (1.9 ± 0.3% shrunken glomeruli in control, 9.8 ± 0.8% in methoxetamine-treated mice (p < 0.0001); 2.9 ± 0.3% tubular cell degeneration in control, 20.4 ± 1.1% in methoxetamine-treated mice (p < 0.0001)). There was a greater density of mononuclear cells in the bladder lamina propria and submucosa in methoxetamine-treated mice (43.0 ± 2.1 per 250 × 250 μm) than controls (7.1 ± 1.2 per 250 × 250 μm), p < 0.001. CD4-positive staining was seen in the bladder submucosa and lamina propria of all methoxetamine-treated mice and muscle-layer of two methoxetamine-treated mice; these changes were not seen in the control mice. There was an increase in sirius-red collagen in the bladder sub-mucosa and muscle-layer in the methoxetamine-treated mice compared with control mice. Discussion. This study has shown that 3 months of daily 30 mg/kg intra-peritoneal methoxetamine results in significant bladder and renal toxicity in mice. Changes in the bladder included inflammatory changes with subsequent fibrosis and changes in the kidney were seen at both a tubular and glomerular level. These changes are similar to those seen in comparable animal models of chronic ketamine administration. Further work is required to determine the time course of the onset of these effects and whether the effects are reversible with methoxetamine cessation.

The Role of the Urothelium and ATP in Mediating Detrusor Smooth Muscle Contractility

OBJECTIVES To examine the contractility of urothelium-intact (+UE) and urothelium-denuded (-UE) rat detrusor strips under adenosine triphosphate (ATP) treatment. Purinergic signaling exists in the bladder but both the inhibitory effect of ATP on detrusor contractions and the function of urothelial ATP are not established. METHODS Detrusor strips were obtained from bladders of young adult rats. Isometric tension from both transverse and longitudinal contractions was measured using a myograph. The muscarinic agonist carbachol (CCh) was used to induce contractions, which were under the influences of different concentrations of ATP. RESULTS In both +UE and -UE strips, 1 mM ATP suppressed CCh-induced contractions. In longitudinal contractions, ATP added to the inhibitory effect of urothelium on CCh responses. Removal of the urothelium, but with exogenous ATP added, recovered the CCh responses to the same level as in +UE strips with no added ATP. Transverse contractions were less susceptible to ATP in the presence of urothelium. CONCLUSIONS We showed that the urothelium and ATP suppressed CCh-induced contractions to a similar extent. The findings suggest an inhibitory role of urothelial ATP in mediating detrusor smooth muscle contractility, which may be impaired in diseased bladders.

Blockade of Voltage-Sensitive K+ Channels Increases Contractility More in Transverse Than in Longitudinal Rat Detrusor Strips

OBJECTIVES To investigate the effects of voltage-sensitive K(+) channel blockade on phasic contractions in transverse (TR) and longitudinal (LO) rat detrusor strips. The contractility of LO detrusor strips has been studied more extensively than that of TR ones, with the latter hardly used in experiments. Phasic contractions, regulated by various types of K(+) channels, are present in normal bladders and are more commonly seen in diseased bladders. METHODS Contractility was measured in TR and LO detrusor strips from young adult rats. Concentration-response curves of tetraethylammonium and 4-aminopyridine (4-AP) were constructed. Both the mean developed tension and the cyclic height of the phasic contractions were determined. RESULTS At > or = 10(-2) mol/L, tetraethylammonium increased the developed tension and amplitude of the phasic contractions without differences between the TR and LO strips. 4-AP also increased the developed tension and amplitude of the phasic contractions but with greater effects in the TR than in the LO strips. At a maximal 4-AP concentration (10(-2) mol/L), the mean developed tension was 52.96% +/- 6.39% in the TR strips and 28.88% +/- 5.91% in the LO strips. The amplitude of the phasic contractions was 11.07% +/- 2.78% for the TR strips and 3.15% +/- 1.31% for the LO strips. CONCLUSIONS Both nonselective K(+) channel blockade by tetraethylammonium and selective voltage-sensitive K(+) channel blockade by 4-AP resulted in increased contractions in LO and TR strips; however, the latter displayed greater effects only with 4-AP treatment. Thus, measuring detrusor contractions in different orientations might reveal heterogeneous responses that are dependent on the pharmacologic stimulus.

Urothelium-dependent and urothelium-independent detrusor contractility mediated by nitric oxide synthase and cyclooxygenase inhibition†‡

The urothelium has been implicated in regulating detrusor smooth muscle contractility but the identity of the putative urothelium‐derived inhibitory factor remains unconfirmed. There was inconclusive evidence on the role of nitric oxide synthase (NOS) and cyclooxygenase (COX) in mediating detrusor contractions. This study examined varying regulation by NOS and COX in transverse and longitudinal carbachol (CCh)‐induced and unstimulated phasic contractions.

The Indispensable Value of Clinical Trials in the Modernization of Traditional Chinese Medicine: 12 Years' Experience at CUHK and Future Perspectives

The last decade has seen a wealth of information reporting the beneficial effects of Chinese herbal medicines. While a lot more studies were done using in vitro and in vivo research platforms, much fewer investigations were conducted according to evidence-based requirements in clinical settings. The Institute of Chinese Medicine at the Chinese University of Hong Kong (CUHK) has had the opportunity to collaborate with clinicians over the years to initiate and conduct dozens of clinical trials investigating and verifying the therapeutic values of Chinese herbs in selected disease conditions. Of the many disorders, we chose to focus on those that are known for their difficulties achieving perfect results with conventional treatment methods. Examples include non-healing ulcers, allergic conditions, degenerative diseases and cancer. Protective effects of the herbs in such chronic diseases as coronary artery disease and osteoporosis were also part of our focus. Even in healthy individuals and those recovering from chemotherapy, Chinese herbs could help with the immune system and were studied in our clinical trials as well. This paper aims to highlight the important findings from these clinical studies while at the same time, stressing the indispensable value of clinical trials in modernizing the use of Chinese herbs in present-day medicine.

Chronic ketamine treatment-induced changes in contractility characteristics of the mouse detrusor

PurposeTo understand bladder contractility changes induced by chronic ketamine treatment, noting the prevalence of its abuse worldwide.MethodsA mouse model of chronic ketamine treatment was used and detrusor strip contractility was measured. Rising and falling phases of contractile responses as well as maximal, average sustained and phasic contractions were measured.ResultsWhile maximal contractility of ketamine-treated strips was identical to the saline controls, the former displayed slower contraction rates under K+-Krebs, carbachol and electrical stimulation. The decay phase of electrically stimulated responses was also slower at most stimulation frequencies in the ketamine-treated strips. Greater sensitivity to varying the strengths of stimuli was observed in the ketamine-treated strips.ConclusionsAltered contractility characteristics of the bladder after chronic ketamine treatment were revealed, which could potentially be useful in the development of improved treatment regimens.

Bladder Contractility Is Mediated by Different K+ Channels in the Urothelium and Detrusor Smooth Muscle

The roles played by K+ channels in the urothelium (UE) and detrusor smooth muscle (DSM) in regulating agonist-induced bladder contraction is not known at present. Thus, the effects in carbachol (CCh)-induced contraction in UE-intact (+UE) and UE-denuded (-UE) rat detrusor strips pretreated with K+-channel blockers were investigated here. The K+-channel blockers used were 4-aminopyridine (4-AP), glibenclamide (Glib), iberiotoxin (IbTx), charybdotoxin (ChTx), and apamin. In the absence of K+-channel blockers, control CCh-induced contractions were more potent in -UE than +UE strips. Treatment with IbTx and apamin resulted in more potent CCh-induced contractions in +UE strips. In -UE strips, CCh potency was increased by ChTx and Glib, but decreased by 4-AP. Different K+ channels in the UE and DSM were thus involved in regulating bladder contractions. Contractile mediatory function of these channels, specific to the UE or DSM, may be potential drug targets in the management of bladder disorders.

Ramipril Improves Oxidative Stress-Related Vascular Endothelial Dysfunction in db/db Mice

Endothelial dysfunction often precedes Type 2 diabetes-associated cardiovascular complications. One important cause of endothelial dysfunction is oxidative stress, which can lead to reduced nitric oxide (NO) bioavailability. In this study, we examined the effects of ramipril (an angiotensin-converting enzyme inhibitor, ACEI) on reactive oxygen species (ROS) production and endothelium-dependent vasodilation using a Type 2 diabetic (db/db) murine model. Plasma concentration of 8-isoprostane ([8-isoP]) was measured and used as an indication of the amount of ROS production. Six weeks of ramipril (10 mg/kg/day) treatment significantly reduced [8-isoP] and improved acetylcholine(ACh)-induced vasodilation in db/db mice without altering responses in wild-type (WT) mice. Responsiveness of smooth muscle cells to NO, assessed by sodium nitroprusside-induced vasodilation, was not different between db/db and WT mice regardless of ramipril or vehicle treatment. Our results suggest that ramipril specifically improved endothelium-dependent vasodilation in Type 2 diabetic mice, possibly by reducing ROS levels.

Variations in carbachol- and ATP-induced contractions of the rat detrusor: effects of gender, mucosa and contractile direction.

PurposeContractile characteristics of the bladder may depend on variables such as gender, mucosa (MU) and direction of the contractions. However, definitive information is not yet available despite earlier studies on the effects of one variable or another. Here, we explored the differences in the rat detrusor attributable to gender, mucosa and contractile direction.MethodsK+, carbachol (CCh) and ATP were used as contractile stimuli on rat detrusor strips with and without MU. Contractility was monitored using a myograph system. Both tonic and phasic contractile activities were analyzed.ResultsMU-independent contractions induced by CCh were more potent in females, an effect specific to the longitudinal direction only. The maximal CCh response was larger also in females when MU was removed, suggesting a stronger MU-independent component in the contraction. The larger area under curves of the females under ATP stimulation showed dependence on MU and contractile direction as well. ATP-induced contractions in the males were affected more by MU in the transverse direction than in the females. Direction- and MU-dependent variability of ATP responses was also observed in the males but not in females.ConclusionsFindings here added new information to the understanding of bladder contractile physiology, providing insights into the quest for better drugs in managing bladder disorders.

Effects of Gegen (Puerariae lobatae Radix) water extract on improving detrusor overactivity in spontaneously hypertensive rats

AIM Ex vivo experiments showed that the water extract of Puerariae lobatae Radix (named Gegen in Chinese) induced detrusor relaxation. The aim of this study was to prove the in vivo efficacy of Gegen on improving detrusor overactivity and its possible synergism with darifenacin (a first-line muscarinic receptor-3 inhibitor) in spontaneously hypertensive rats (SHR), a rat model exhibiting symptoms of detrusor overactivity. METHOD After daily oral administration of Gegen 30 (Gegen, 30mg/kg); Gegen 300 (Gegen, 300mg/kg); Low_Dar (darifenacin, 3mg/kg); High_Dar (darifenacin, 30mg/kg) Low_Dar+Gegen 30 or High_Dar+Gegen 30 for 3 weeks, bladder detrusor strips of the rats were isolated and assessed with different stimulators for the measurement of tonic and phasic contractile activities (including phasic amplitude and frequency). Modes of stimulation included the use of carbachol, isoprenaline and electrical field stimulation (EFS). RESULTS All drug treatments significantly reduced carbachol-stimulated tonic contractile activities, but did not change the phasic amplitude. Meanwhile, the treatments with Gegen 300; Low_Dar; Low_Dar+Gegen 30; and High_Dar+Gegen 30 decreased carbachol-stimulated phasic frequency. Gegen 300 and Low_Dar+Gegen 30 showed stronger potency on lowering EFS-induced responses. Under isoprenaline-induced relaxation, only Gegen 300 significantly enhanced this relaxation by decreasing tonic contraction; Gegen 300; Low_Dar; Low_Dar+Gegen 30; and High_Dar+Gegen 30 increased the reduction of phasic frequency, but all treatment did not alter their phasic amplitude. Combination Index (CI) showed that the combination with Low_Dar and Gegen 30 had very strong synergism (CI <0.1) on inhibiting EFS-induced contractile response. CONCLUSION Gegen improved detrusor overactivity through neurogenic and anti-muscarinic mechanisms. Gegen and darifenacin together attained synergism for detrusor overactivity treatment via the neurogenic pathway.