Organ bath experiments are a key technology to assess contractility of smooth muscle. Despite efforts to standardize tissue specimen sizes, they vary to a certain degree. As it appears obvious that a larger piece of tissue should develop greater force, most investigators normalize contraction data for specimen size. However, they lack agreement which parameter should be used as denominator for normalization. A pre-planned analysis of data from a recent study was used to compare denominators used for normalization, i.e., weight, length, and cross-sectional area. To increase robustness, we compared force with denominator in correlation analysis and also coefficient of variation with different denominators. This was done concomitantly with urinary bladder strips and aortic rings and with multiple contractile stimuli. Our urinary bladder data show that normalization for strip weight yielded the tightest but still only moderate correlation (e.g., r2 = 0.3582 for peak carbachol responses based on 188 strips). In aorta, correlations were even weaker (e.g., r2 = 0.0511 for plateau phenylephrine responses normalized for weight based on 200 rings). Normalization for strip size is less effective in reducing data variability than previously assumed; the normalization denominator of choice must be identified separately for each preparation.
This is a preview of subscription content, log in to check access.
Buy single article
Instant access to the full article PDF.
Tax calculation will be finalised during checkout.
Subscribe to journal
Immediate online access to all issues from 2019. Subscription will auto renew annually.
Tax calculation will be finalised during checkout.
Abebe W, Harris KH, MacLeod KM (1990) Enhanced contractile responses of arteries from diabetic rats to α1-adrenoceptor stimulation in the absence and presence of extracellular calcium. J Cardiovasc Pharmacol 16:239–248
Braverman AS, Ruggieri MR Sr (2003) Hypertrophy changes the muscarinic receptor subtype mediating bladder contraction from M3 toward M2. Am J Phys 285:R701–R708
Cameron NE, Cotter MA (1992) Impaired contraction and relaxation in aorta from streptozotocin-diabetic rats: role of polyol pathway. Diabetologia 35:1011–1019
Conklin DJ, Boor PJ (1998) Allylamine cardiovascular toxicity: evidence for aberrant vasoreactivity in rats. Toxicol Appl Pharmacol 148:245–251
Conklin DJ, Cowley HR, Wiechmann RJ, Johnson GH, Trent MB, Boor PJ (2004) Vasoactive effects of methylamine in isolated human blood vessels: role of semicarbazide-sensitive amine oxidase, formaldehyde, and hydrogen peroxide. Am J Phys 286:H667–H676
Conklin DJ, Haberzettl P, Prough RA, Bhatnagar A (2009) Glutathione-S-transferase P protects against endothelial dysfunction induced by exposure to tobacco smoke. Am J Phys 296:H1586–H1597
Frazier EP, Braverman AS, Peters SLM, Michel MC, Ruggieri MR Sr (2007) Does phospholipase C mediate muscarinic receptor-induced rat urinary bladder contraction? J Pharmacol Exp Ther 322:998–1002
Fujishige A, Takahashi K, Tsuchiya T (2002) Altered mechanical properties in smooth muscle of mice with a mutated calponin locus. Zool Sci 19:167–174
Halsey LG, Curran-Everett D, Vowler SL, Drummond GB (2015) The fickle P value generates irreproducible results. Nat Med 12:179–185
Jin L, Lipinski A, Conklin DJ (2018) A simple method for normalization of aortic contractility. J Vasc Res 55:177–186
Kershaw JDB, Misfield M, Sievers H-H, Yacoub MH, Chester AH (2004) Specific regional and directional contractile responses of aortic cup tissue. J Heart Valve Dis 13:798.803
Kilkenny C, Browne W, Cuthill IC, Emerson M, Altman DG (2010) Animal research: reporting in vivo experiments: the ARRIVE guidelines. Br J Pharmacol 160:1577–1579
Klausner AP, Rourke KF, Miner AS, Ratz PH (2009) Potentiation of carbachol-induced detrusor smooth muscle contractions by ß-adrenoceptor activation. Eur J Pharmacol 606:191–198
Kories C, Czyborra C, Fetscher C, Schneider T, Krege S, Michel MC (2003) Gender comparison of muscarinic receptor expression and function in rat and human urinary bladder: differential regulation of M2 and M3? Naunyn Schmiedeberg's Arch Pharmacol 367:524–531
Kristek F, Cacanyiova S, Gerova M (2009) Hypotrophic effect of long-term neuronal NO-synthase inhibition on heart and conduit arteries of the Wistar rats. J Physiol Pharmacol 60:21–27
Kunert MP, Dwinell MR, Lombard JH (2010) Vascular responses in aortic rings of a consomic rat panel derived from the fawn hooded hypertensive strain. Physiol Genomics 42A:244–258
Longhurst PA, Kang J, Wein AJ, Levin RM (1990) Length-tension relationship of urinary bladder strips from streptozotocin-diabetic rats. Pharmacology 40:110–121
Meini S, Lecci A, Cucchi P, Catalioto RM, Criscuoli M, Maggi CA (1998) Inflammation modifies the role of cyclooxygenases in the contractile responses of the rat detrusor smooth muscle to kinin agonists. J Pharmacol Exp Ther 287:137–143
Michel MC (2014) Do ß-adrenoceptor agonists induce homologous or heterologous desensitization in rat urinary bladder? Naunyn Schmiedeberg's Arch Pharmacol 387:215–224
Michel-Reher MB, Michel MC (2015) Regulation of GAPDH expression by treatment with the ß-adrenoceptor agonist isoprenaline - is GAPDH a suitable loading control in immunoblot experiments? Naunyn Schmiedeberg's Arch Pharmacol 388:1119–1120
Ozcelikay AT, Tay A, Güner S, Tasyaran V, Yildizoglu-Ari N, Dincer UD, Altan VM (2000) Reversal effects of L-arginine treatment on blood pressure and vascular responsiveness of streptozotocin-diabetic rats. Pharmacol Res 41:201–209
Paro M, Italiano G, Travagli RA, Petrelli L, Zanoni R, Prosdocimi M, Fiori MG (1990) Cystometric changes in alloxan diabetic rat: evidence for functional and structural correlates of diabetic autonomic neuropathy. J Auton Nerv Syst 30:1–11
Pieper GM, Langenstroer P, Siebeneich W (1997) Diabetic-induced endothelial dysfunction in rat aorta: role of hydroxyl radicals. Cardiovasc Res 34:145–156
Sand C, Michel MC (2014) Bradykinin contracts rat urinary blader largely independent of phospholipase C. J Pharmacol Exp Ther 348:25–31
Schneider T, Fetscher C, Krege S, Michel MC (2004a) Signal transduction underlying carbachol-induced contraction of human urinary bladder. J Pharmacol Exp Ther 309:1148–1153
Schneider T, Hein P, Michel MC (2004b) Signal transduction underlying carbachol-induced contraction of rat urinary bladder. I. Phospholipases and Ca2+ sources. J Pharmacol Exp Ther 308:47–53
Schneider T, Hein P, Bai J, Michel MC (2005a) A role for muscarinic receptors or rho-kinase in hypertension associated rat bladder dysfunction? J Urol 173:2178–2181
Schneider T, Hein P, Michel-Reher M, Michel MC (2005b) Effects of ageing on muscarinic receptor subtypes and function in rat urinary bladder. Naunyn Schmiedeberg's Arch Pharmacol 372:71–78
Schneider T, Fetscher C, Michel MC (2011) Human urinary bladder strip relaxation by the ß-adrenoceptor agonist isoprenaline: methodological considerations and effects of gender and age. Front Pharmacol 2:11
Stevens LA, Sellers DJ, McKay NG, Chapple CR, Chess-Williams R (2006) Muscarinic receptor function, density and G-protein coupling in the overactive diabetic rat bladder. Auton Autacoid Pharmacol 26:303–309
Török J, Zemancikova A, Kocianova Z (2016) Interaction of perivascular adipose tissue and sympathetic nerves in arteries from normotensive and hypertensive rats. Phys Rep 65(Suppl. 3):S391–S399
Weber LP, Chow WL, Abebe W, MacLeod KM (1996) Enhanced contractile responses of arteries from streptozotocin diabetic rats to sodium fluoride. Br J Pharmacol 118:115–122
Wyse DG (1980) On the “normalization” of active developed force of isolated helical strips of muscular and elastic arteries for variation in wall thickness. J Pharmacol Methods 4:313–326
Yang Q, Fujii W, Kaji N, Kakuta S, Kada K, Kuwahara M, Tsubone H, Ozaki H, Hori M (2018) The essential role of phospho-T38 CPI-17 in the maintenance of physiological blood pressure using genetically modified mice. FASEB J 32:2095–2109
Yesilyurt ZE, Erdogan BR, Karaomerlioglu I, Müderrisoglu AE, Michel MC, Arioglu Inan E (2019) Urinary bladder weight and function in a rat model of mild hyperglycemia and its treatment with dapagliflozin. Front Pharmacol 10:911
This study was supported by in part by grants from Ankara University (BAP-16L0237006), the Scientific and Technological Research Council of Turkey (TUBITAK SBAG-115S564), Deutsche Forschungsgemeinschaft (Mi 294/8-1), and the Innovative Medicines Initiative 2 Joint Undertaking (grant agreement no. 777364); this Joint Undertaking receives support from the European Union’s Horizon 2020 research and innovation program and EFPIA). BRE is a PhD student supported by Scientific and Technical Research Council of Turkey (TUBITAK-2211/A) and by a grant from ERASMUS+ Program of the European Union.
The study protocol had been approved by the animal welfare committee of Ankara University (permit 2015-4-82, 2017-10-92) and was in line with NIH Guidelines for Care and Use of Laboratory Animals.
Conflict of interest
The authors report no conflict of interest related to this paper.
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Electronic supplementary material
About this article
Cite this article
Erdogan, B.R., Karaomerlioglu, I., Yesilyurt, Z.E. et al. Normalization of organ bath contraction data for tissue specimen size: does one approach fit all?. Naunyn-Schmiedeberg's Arch Pharmacol 393, 243–251 (2020). https://doi.org/10.1007/s00210-019-01727-x
- Urinary bladder
- Cross-sectional area