Functional and molecular characterization of endothelium-dependent and endothelium-independent relaxant pathways in uterine artery of non-pregnant buffaloes
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Present study was undertaken to unravel the endothelium-dependent and endothelium-independent relaxant pathways in uterine artery of non-pregnant buffaloes. Isometric tension of arterial rings was recorded using data acquisition system based polyphysiograph. Acetylcholine (ACh) produced endothelium-dependent vasorelaxation by releasing nitric oxide (NO), and inhibition of nitric oxide synthase (NOS) by L-NAME (300 μM) significantly (P < 0.05) reduced the NO release and thereby the vasorelaxant effect of ACh. However, L-NMMA, another NOS inhibitor, and PTIO, a NO scavenger, did not have any additional inhibitory effect on NO and ACh-induced vasorelaxation. Cyclooxygenase (COX) inhibitor (indomethacin) alone did not have any inhibitory action on vasorelaxant response to ACh; however, simultaneous inhibition of COX and NOS enzymes significantly (P < 0.05) attenuated the relaxant response indicating the concurrent release of these two mediators in regulating ACh-induced relaxation. Besides NOS and COX-derived metabolites (EDRF), small (SKCa) and intermediate (IKCa) conductance K+ channels being the members of EDHF play predominant role in mediating ACh-induced vasorelaxation. Using different molecular tools, existence of eNOS, COX-1, and,IKCa in the endothelium, BKCa in vascular smooth muscle, and SKCa in both endothelium and vascular smooth muscle was demonstrated in buffalo uterine artery. Gene sequencing of COX-1 and SKCa genes in uterine artery of buffaloes showed more than 97% structural similarity with ovine (Ovis aries), caprine (Capra hircus), and Indian cow (Bos indicus). Endothelium-independent nitrovasodilator, sodium nitroprusside (SNP), produced vasorelaxation which was sensitive to blockade by soluble guanylate cyclase (sGC) inhibitor (ODQ), thus suggesting the important role of cGMP/PKG pathways in uterine vasorelaxation in buffaloes. Taken together, it is concluded that both endothelium-dependent (EDHF and EDRF) and endothelium-independent (sGC-cGMP) relaxant pathways are present in uterine arteries of non-pregnant buffaloes, and they differently contribute to vasorelaxation during non-pregnant state.
KeywordsNitric oxide Cyclooxygenase K+ channels Uterine artery Non-pregnant Buffalo
UPN, AS, and VS conducted functional experiments; UPN, SC, PK, SVN, and KKN conducted molecular experiments; SC and SKG designed the research; UPN, SC and SB analyzed the data, and SC and SKG wrote the manuscript.
Research work presented in this manuscript was supported by Indian Council of Agricultural Research, New Delhi, India under Niche Area of Excellence Programme (Grant No. 10 (10)/2012-EPD dated 23 March 2012) to Department of Veterinary Pharmacology and Toxicology, DUVASU, Mathura, India. Financial assistance by ICAR is thankfully acknowledged. The first author was also awarded the INSPIRE fellowship from Department of Science & Technology (DST-INDIA) via Grant No. DST/INSPIRE Fellowship/2014/IF140997.
Compliance with ethical standards
Conduct of experiments on uterus, which are non-edible and thrown away by the slaughterers, was collected from the slaughterhouse or butcher’s shops, and these are not governed by the “Guidelines of the Committee for the Purpose of Control and Supervision of Experiments on Animals” of the Government of India as resolved by the Institute Animal Ethics Committee in its meeting held on 16 September 2016 (Institutional Animal Ethics Committee; Approval No. 110/IAEC/16/40/2).
Conflict of interest
The authors declare that they have no conflict of interest.
- Ali A, Abdel‐Razek AK, Abdel‐Ghaffar S, Glatzel PS (2003) Ovarian follicular dynamics in buffalo cows (Bubalus bubalis). Reprod Domest Anim 38(3):214–218Google Scholar
- Bird IM, Zhang L, Magness RR (2003) Possible mechanisms underlying pregnancy-induced changes in uterine artery endothelial function. Am J Phys Regul Integr Comp Phys 284(2):R245–R258Google Scholar
- Boura ALA, Walters WAW, Read MA, Leitch IM (1994) Autacoids and control of human placental blood flow. Invited review Clin Exp Pharmacol Physiol 21:737–748Google Scholar
- Choudhury S, Kannan K, Pule Addison M, Darzi SA, Singh V, Singh TU, Thangamalai R, Dash JR, Parida S, Debroy B, Paul A, Mishra SK (2015) Combined treatment with atorvastatin and imipenem improves survival and vascular functions in mouse model of sepsis. Vasc Pharmacol 71:139–150CrossRefGoogle Scholar
- Eckman DM, Gupta R, Rosenfeld CR, Morgan TM, Charles SM, Mertz H, Moore LG (2012) Pregnancy increases myometrial artery myogenic tone via NOS- or COX-independent mechanisms. Am J Phys Regul Integr Comp Phys 303(4):R368–R375Google Scholar
- Fouty B, Komalavilas P, Muramatsu M, Cohen A, McMurtry IF, Lincoln TM, Rodman DM (1998) Protein kinase G is not essential to NO-cGMP modulation of basal tone in rat pulmonary circulation. Am J Phys 274(2 Pt 2):H672–H678Google Scholar
- Gangula PR, Zhao H, Supowit S, Wimalawansa S, DiPette D, Yallampalli C (1999) Pregnancy and steroid hormones enhance the vasodilation responses to CGRP in rats. Am J Phys 276(1 pt 2):H284–H288Google Scholar
- Gebremedhin D, Kaldunski M, Jacobs ER, Harder DR, Roman RJ (1996) Co-existence of two types of calcium activated potassium channels in rat renal arterioles. Am J Phys 270:F69–F81Google Scholar
- Kovitz K, Aleskowitch TD, Sylvester JT, Flavahan NA (1993) Endothelium derived contracting and relaxing factors contribute to hypoxic responses of pulmonary arteries. Am J Phys 256:H1139–H1148Google Scholar
- Ledoux J, Werner ME, Brayden JE, Nelson MT (2006) Calcium-activated potassium channels and the regulation of vascular tone. Physiology (Bethesda) 21:69–78Google Scholar
- Lugnier C, Komas N (1993) Modulation of vascular cyclic nucleotide phosphodiesterases by cyclic GMP: role in vasodilatation. Eur Heart J 14(suppl. I):141–148Google Scholar
- Lunell NO, Nylund LE, Lewander R, Sarby B (1982) Uteroplacental blood flow in pre-eclampsia measurements with indium-113m and a computer-linked gamma camera. Clin Hypertens B 1:105–117Google Scholar
- Magness RR (1991) National Institutes of Health symposium. Endothelium-derived vasoactive substances and uterine blood vessels. Semin Perinatol 15:68–78Google Scholar
- Scotland RS, Madhani M, Chauhan S, Moncada S, Andresen J, Nilsson H, Hobbs AJ, Ahluwalia A (2005) Investigation of vascular responses in endothelial nitric oxide synthase/cyclooxygenase-1 double-knockout mice: key role for endothelium-derived hyperpolarizing factor in the regulation of blood pressure in vivo. Circulation. 111(6):796–803CrossRefGoogle Scholar
- Senadheera S, Bertrand PP, Grayson TH, Leader L, Murphy TV, Sandow SL (2013) Pregnancy-induced remodelling and enhanced endothelium-derived hyperpolarization-type vasodilator activity in rat uterine radial artery: transient receptor potential vanilloid type 4 channels, caveolae and myoendothelial gap junctions. J Anat 223(6):677–686CrossRefGoogle Scholar
- Senadheera S, Kim Y, Grayson TH, Toemoe S, Kochukov MY, Abramowitz J, Housley GD, Bertrand RL, Chadha PS, Bertrand PP, Murphy TV, Tare M, Birnbaumer L, Marrelli SP, Sandow SL (2012) Transient receptor potential canonical type 3 channels facilitate endothelium-derived hyperpolarization mediated resistance artery vasodilator activity. Cardiovasc Res 95:439–447CrossRefGoogle Scholar
- Sladek SM, Magness RR, Conrad KP (1997) Nitric oxide and pregnancy. Am J Phys Regul Integr Comp Phys 272:R441–R463Google Scholar
- Sukumaran SV, Singh TU, Parida S, Reddy CEN, Thangamalai R, Kandasamy K, Singh V, Mishra SK (2013) TRPV4 channel activation leads to endothelium-dependent relaxation mediated by nitric oxide and endothelium-derived hyperpolarizing factor in rat pulmonary artery. Pharmacol Res 78:18–27CrossRefGoogle Scholar
- Vanhoutte PM (1996) Endothelium-derived hyperpolarizing factor, vol XXI. Harwood academic publishers, Amsterdam, p 338Google Scholar