Characterization of the control of intracellular [Ca2+] and the contractile phenotype of cultured human detrusor smooth muscle cells.
OBJECTIVE
We measured the functional properties of cultured human detrusor myocytes with respect to their ability to regulate their intracellular [Ca2+] and generate force in collagen matrices.


METHODS
Human detrusor biopsies were dissociated into single cells by collagenase treatment and used immediately or cultured in D-valine medium and subsequently used after culture trypsinization. Intracellular [Ca2+] was measured in Fura-2 loaded myocytes. Cell force development was measured by incorporating cells into a collagen gel and attaching it to an isometric strain gauge.


RESULTS
Carbachol was equally effective in generating Ca transients in freshly isolated and cultured cells. Carbachol potency (pEC50) and the magnitude of Ca2+ transients were similar. Adenosine triphosphate potency was decreased in cultured cells and Ca2+ transients showed properties consistent with a purinoceptor shift from a purinergic subtype. Temporal restitution of Ca2+ transients was similar in the 2 groups, indicative of retained intracellular Ca2+ stores in cultured cells. Cultured cells (approximately 10(6)) embedded in collagen gel generated a force about 10 times greater than that generated by gel alone. The cell dependent force could be further increased by adding carbachol.


CONCLUSIONS
Cultured cells retain the ability to generate agonist induced intracellular Ca2+ transients. There was no evidence that the cell culture altered the properties of muscarinic receptors, although purinoceptor mediated properties were altered. Restitution experiments indicated that functional intracellular Ca2+ stores were retained in cultured cells. Cultured cells also retained a contractile phenotype, especially in response to carbachol. The magnitude of force was attenuated, which may be a function of the biomechanical properties of the gel used to embed the cells.
Wood
D N
DN
Institute of Urology and Nephrology, and Tissue Repair and Engineering Centre, London, United Kingdom. drdanwood@mac.com
Brown
R A
RA
Fry
C H
CH
D002118:Calcium / Q000502:physiology*; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D018679:Cholinergic Agonists / Q000494:pharmacology; D006801:Humans; D009119:Muscle Contraction / Q000502:physiology*; D009130:Muscle, Smooth / Q000502:physiology*; D001743:Urinary Bladder / Q000502:physiology
D018679:Cholinergic Agonists; D002217:Carbachol; D002118:Calcium