Bitopic Sphingosine 1-Phosphate Receptor 3 (S1P3) Antagonist Rescue from Complete Heart Block: Pharmacological and Genetic Evidence for Direct S1P3 Regulation of Mouse Cardiac Conduction.
The molecular pharmacology of the G protein-coupled receptors for sphingosine 1-phosphate (S1P) provides important insight into established and new therapeutic targets. A new, potent bitopic S1P3 antagonist, SPM-354, with in vivo activity, has been used, together with S1P3-knockin and S1P3-knockout mice to define the spatial and functional properties of S1P3 in regulating cardiac conduction. We show that S1P3 is a key direct regulator of cardiac rhythm both in vivo and in isolated perfused hearts. 2-Amino-2-[2-(4-octylphenyl)ethyl]propane-1,3-diol in vivo and S1P in isolated hearts induced a spectrum of cardiac effects, ranging from sinus bradycardia to complete heart block, as measured by a surface electrocardiogram in anesthetized mice and in volume-conducted Langendorff preparations. The agonist effects on complete heart block are absent in S1P3-knockout mice and are reversed in wild-type mice with SPM-354, as characterized and described here. Homologous knockin of S1P3-mCherry is fully functional pharmacologically and is strongly expressed by immunohistochemistry confocal microscopy in Hyperpolarization Activated Cyclic Nucleotide Gated Potassium Channel 4 (HCN4)-positive atrioventricular node and His-Purkinje fibers, with relative less expression in the HCN4-positive sinoatrial node. In Langendorff studies, at constant pressure, SPM-354 restored sinus rhythm in S1P-induced complete heart block and fully reversed S1P-mediated bradycardia. S1P3 distribution and function in the mouse ventricular cardiac conduction system suggest a direct mechanism for heart block risk that should be further studied in humans. A richer understanding of receptor and ligand usage in the pacemaker cells of the cardiac system is likely to be useful in understanding ventricular conduction in health, disease, and pharmacology.
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
M Germana
MG
Sanna
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Kevin P
KP
Vincent
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Emanuela
E
Repetto
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Nhan
N
Nguyen
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Steven J
SJ
Brown
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Lusine
L
Abgaryan
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Sean W
SW
Riley
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Nora B
NB
Leaf
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Stuart M
SM
Cahalan
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
William B
WB
Kiosses
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Yasushi
Y
Kohno
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Joan Heller
JH
Brown
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.).
Andrew D
AD
McCulloch
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.) hrosen@scripps.edu gonzalp@scripps.edu.
Hugh
H
Rosen
Departments of Chemical Physiology (M.G.S., E.R., N.N., H.R., P.J.G.-C.), Immunology (N.B.L.), and Molecular and Cellular Neuroscience (S.M.C.), Scripps Research Institute Molecular Screening Center (S.J.B., L.A., S.W.R.), Microscopy Core (W.B.K.), Scripps Research Institute, La Jolla, California; Kyorin Pharmaceutical Company, LTD, Tokyo, Japan (Y.K.); and Departments of Bioengineering (A.D.M., K.P.V.) and Pharmacology, University of California, San Diego, California (J.H.B.) hrosen@scripps.edu gonzalp@scripps.edu.
Pedro J
PJ
Gonzalez-Cabrera
D000818:Animals; D002316:Cardiotonic Agents / Q000494:pharmacology / Q000627:therapeutic use; D006327:Heart Block / Q000188:drug therapy* / Q000235:genetics* / Q000503:physiopathology; D006339:Heart Rate / Q000187:drug effects* / Q000502:physiology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D049349:Receptors, Lysosphingolipid / Q000037:antagonists & inhibitors* / Q000235:genetics*; D000081025:Sphingosine-1-Phosphate Receptors
D002316:Cardiotonic Agents; D049349:Receptors, Lysosphingolipid; C501285:S1pr3 protein, mouse; D000081025:Sphingosine-1-Phosphate Receptors