Abstract Body: INTRODUCTION
Histamine (H) binds to its cognate G-protein coupled receptors (GPCRs) H1R and H2R to regulate cardiac function. The activation of H1R inhibits, while that of H2R stimulates, cAMP production. These cAMP pools are hydrolyzed by phosphodiesterases (PDEs) 1, 3, and 4 in the human heart. All three PDE inhibitors are in clinical use or under investigation, including for heart failure. However, how PDEs dynamically work with these receptors to alter heart function is unknown.

METHODS
We first determined which PDEs hydrolyze the Gs-coupled H2R cAMP pool. Using a cAMP biosensor, we performed Forster resonance energy transfer experiments in guinea pig adult ventricular myocytes. Specific GPCRs or PDEs were pharmacologically targeted. To test the functional effects of targeting specific GPCR/PDE pairs, cell sarcomere shortening and Ca2+ transients were recorded in paced myocytes.

RESULTS
As already shown in rat atrial preparations, we hypothesized that PDE3 would regulate H2R-associated cAMP pool in guinea pig ventricular myocytes. FRET experiments confirmed a strong role for PDE3 and lesser but significant role for PDE4 to regulate H2R-associated cAMP pool. PDE1 regulated the cAMP pool associated with adenosine2A receptor (A2AR), but neither A2BR nor H2R. GPCR/PDE pairings that synergistically regulated cAMP pool also exerted positive inotropic and lusitropic changes in cell sarcomere shortening. Unexpectedly, we found that the synergistic regulation of cAMP was lost when cells are treated with a PDE inhibitor and then a GPCR agonist. Prior research suggested heterodimerizations or interactions between GPCRs can switch whether Gs or Gi protein binds to adenylyl cyclase. We next hypothesized that HR may override AR signaling in this manner. As expected, blocking H1R reduced the effects of H2R-mediated cAMP production. First treating cells with H2R agonist blocked A2AR- and A2BR-mediated cAMP production. When reversed, A2BR but not A2AR stimulation amplified the effects of H2R agonist.

CONCLUSION
GPCR and PDE interactions stratify cardiovascular effects of histamine: 1) PDEs 3 and 4 regulate H2R-associated cAMP pools, 2) PDE1 does this with A2AR, 3) H2R activation blocks A2A and A2BR signaling, and 4) A2BR priming increases H2R-mediated cAMP production.