5-HT4 receptor-mediated vasorelaxation occurs through a cyclic mononucleotide-dependent mechanism in the isolated bovine lateral saphenous vein [abstract]

Authors: TROTTA, RONALD - University Of Kentucky; HARMON, DAVID - University Of Kentucky; Klotz, James

Submitted to: American Society of Animal Science Annual Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 4/11/2024
Publication Date: 9/18/2024
Citation: Trotta, R.J., Harmon, D.L., Klotz, J.L. 2024. 5-HT4 receptor-mediated vasorelaxation occurs through a cyclic mononucleotide-dependent mechanism in the isolated bovine lateral saphenous vein [abstract]. In: Journal of Animal Science. American Society of Animal Science Annual Meeting, Calgary, Canada July 21-25, 2024. 102 S3:268-269.

Interpretive Summary:

Technical Abstract: Serotonin (5-HT)-mediated vasorelaxation of the isolated bovine lateral saphenous vein occurs primarily through 5-HT4 receptor activation. Vasorelaxation responses mediated through 5-HT4 have only been documented to occur in cattle and sheep and thus, may represent a novel vasorelaxation mechanism that is specific to ruminants. In tissues from other species, the 5-HT4 receptor and its associated G protein couple adenylate cyclase. However, the mechanisms contributing to 5-HT4-mediated vasorelaxation in vascular tissue remain undefined. To better understand the mechanisms of 5-HT4-mediated vasorelaxation, lateral saphenous veins from cattle (n=4) were collected and assessed for vasoactivity in response to increasing concentrations of a selective 5-HT4 receptor agonist (BIMU-8) in the absence and presence of selective inhibitors of downstream proteins involved with cyclic mononucleotide-mediated signaling. Isolated vessel segments were suspended in chambers of a multi-myograph containing 5 mL of continuously oxygenated Krebs-Henseleit buffer and equilibrated to 1 g tension for 90 min. After the equilibration period, vessels were exposed to dimethyl sulfoxide (DMSO; vehicle control) or selective inhibitors of downstream protein targets dissolved in DMSO (1'10-5 M) for 5 min. Vessels were then pre-contracted with 1'10-4 M phenylephrine for 10 min to achieve a sustained increase in vessel tension. After the 10-min pre-contraction period, vessels were exposed to increasing concentrations (1'10-9 to 1'10-4) of BIMU-8 non-cumulatively. Vasoactive response data were normalized as a percentage of the maximum contractile response induced by the phenylephrine pre-contraction. In the absence of selective inhibitors (control), BIMU-8 induced (P<0.01) dose-dependent relaxations (mean -log EC50 = 6.30 M) with a 112% increase in maximal vasorelaxation activity occurring at 1'10-4 M. In the presence of the selective inhibitors for adenylate cyclase (NKY80) or guanylate cyclase (NS2028), 66% and 86% of the vasorelaxation responses to BIMU-8 were attenuated (P<0.01), respectively. In the presence of the selective inhibitors for protein kinase A [protein kinase A inhibitor fragment (6-22) amide] or protein kinase G (Rp-8-bromo-PET-Cyclic GMPS), 66% and 97% of the maximal vasorelaxation responses to BIMU-8 were attenuated (P<0.01), respectively. In the presence of non-selective inhibitors for Ca2+ channels and K+ channels, vasorelaxation responses to BIMU-8 were attenuated (P<0.01). In the presence of selective inhibitors for the protein phosphatase 1 and 2A subunits of myosin light chain (calyculin A) and heat shock protein synthesis (quercetin), vasorelaxation responses to BIMU-8 were attenuated (P<0.01). The findings of the current study demonstrate that 5-HT4-receptor mediated vasorelaxation occurs through a cyclic mononucleotide-dependent mechanism involving cross-activation of protein kinase A and G signaling. Downstream proteins involved in the signaling cascade leading to vasorelaxation include Ca2+ channels, K+ channels, myosin light chain phosphatase, and heat shock proteins. More research is needed to fully describe the cellular and molecular mechanisms responsible for 5-HT4 receptor-mediated vasorelaxation.