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Changes in muscarinic receptors in the toad urothelial cell line TBM-54 following acrolein treatment

Journal article
Authors Daniel Giglio
Michael Andersson
Patrik Aronsson
Dick Delbro
Börje Haraldsson
Gunnar Tobin
Published in Clinical and Experimental Pharmacology and Physiology
Volume 35
Issue 2
Pages 217-222
ISSN 0305-1870
Publication year 2008
Published at Institute of Neuroscience and Physiology, Department of Pharmacology
Institute of Medicine, Department of Molecular and Clinical Medicine
Institute of Clinical Sciences
Pages 217-222
Language en
Keywords urothelium, muscarinic, receptor, cyclophosphamide, cystitis
Subject categories Pharmacology


1. In cyclophosphamide-induced cystitis in the rat, cholinergic function of the bladder and muscarinic receptor expression are altered. In the present study, we investigated whether the toad urothelial cell line TBM-54 expresses functional muscarinic receptors and whether changes in muscarinic receptors can be induced in vitro by treating cells with acrolein, a metabolite of cyclophosphamide causing cystitis. 2. The occurrence of muscarinic receptors on cells was assessed by microphysiometry, a method analysing receptor function by measuring changes in the extracellular acidity rate (ECAR) in response to receptor stimulation. 3. Challenging untreated cells with the muscarinic receptor agonist carbachol gave rise to a concentration-dependent increase in changes in ECAR, with a maximal response at 1 mmol/L carbachol of 51 +/- 6%. Pre-incubating cells with different muscarinic receptor antagonists (i.e. pirenzepine (M(1) receptor selective), methoctramine (M(2)/M(4) receptor selective) and 4-diphenylacetoxy-N-methylpiperidine methobromide (4-DAMP; M(3)/M(1)/M(5) receptor selective)), gave rise to a concentration-dependent decrease in the effects of carbachol (0.5 mmol/L) on changes in ECAR. 4. Western blot analysis was used to determine the expression of all muscarinic receptor subtypes (M(1)-M(5)) by the cell line. Following acrolein treatment, cells were markedly less sensitive to carbachol and the expression of muscarinic M(2) receptors was decreased, whereas the expression of muscarinic M(3) receptors was increased. 5. In conclusion, the urothelial cell line TBM-54 expresses functional muscarinic receptors and exposure to acrolein leads to a modulation in the expression of muscarinic receptors. Consequently, acrolein may have direct effects on muscarinic receptor function and expression that contribute to the pathogenesis of cyclophosphamide-induced cystitis.

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