phosphatidylinositol-4-phosphate and Cardiomegaly

Increased adrenergic tone resulting from cardiovascular stress leads to development of heart failure, in part, through chronic stimulation of β1 adrenergic receptors (βARs) on cardiac myocytes. Blocking these receptors is part of the basis for β-blocker therapy for heart failure. Recent data demonstrate that G protein-coupled receptors (GPCRs), including βARs, are activated intracellularly, although the biological significance is unclear. Here we investigated the functional role of Golgi βARs in rat cardiac myocytes and found they activate Golgi localized, prohypertrophic, phosphoinositide hydrolysis, that is not accessed by cell surface βAR stimulation. This pathway is accessed by the physiological neurotransmitter norepinephrine (NE) via an Oct3 organic cation transporter. Blockade of Oct3 or specific blockade of Golgi resident β1ARs prevents NE dependent cardiac myocyte hypertrophy. This clearly defines a pathway activated by internal GPCRs in a biologically relevant cell type and has implications for development of more efficacious β-blocker therapies.

Topics: Animals; Cardiomegaly; Golgi Apparatus; Guanine Nucleotide Exchange Factors; Hydrolysis; Myocytes, Cardiac; Phosphatidylinositol Phosphates; Phosphoinositide Phospholipase C; Rats; Receptors, Adrenergic, beta-1

In cardiac myocytes activation of an exchange factor activated by cAMP (Epac) leads to activation of phospholipase Cε (PLCε)-dependent hydrolysis of phosphatidylinositol 4-phosphate (PI4P) in the Golgi apparatus a process critical for development of cardiac hypertrophy. Here we show that β-adrenergic receptor (βAR) stimulation does not stimulate this pathway in the presence of the broad spectrum phosphodiesterase (PDE) inhibitor IBMX, but selective PDE3 inhibition revealed βAR-dependent PI4P depletion. On the other hand, selective inhibition of PDE2 or PDE9A blocked endothelin-1 (ET-1) and cAMP-dependent PI4P hydrolysis by PLCε. Direct activation of protein kinase A (PKA), protein kinase G (PKG), or the atrial natriuretic factor (ANF) receptor abolished PI4P hydrolysis in response to multiple upstream stimuli. These results reveal distinct pools of cyclic nucleotides that either inhibit PLCε at the Golgi through PKA/PKG, or activate PLCε at the Golgi through Epac. These data together reveal a new mechanism by which ANF and selective PDE inhibitors can protect against cardiac hypertrophy.

Topics: 1-Methyl-3-isobutylxanthine; A Kinase Anchor Proteins; Animals; Atrial Natriuretic Factor; Cardiomegaly; Cardiomyopathy, Hypertrophic; Cell Compartmentation; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Golgi Apparatus; Humans; Myocytes, Cardiac; Nucleotides; Phosphatidylinositol Phosphates; Phosphoinositide Phospholipase C; Phosphoric Diester Hydrolases; Rats; Rats, Sprague-Dawley; Receptors, Adrenergic, beta; Signal Transduction

We recently identified a novel GPCR-dependent pathway for regulation of cardiac hypertrophy that depends on Golgi phosphatidylinositol 4-phosphate (PI4P) hydrolysis by a specific isoform of phospholipase C (PLC), PLCε, at the nuclear envelope. How stimuli are transmitted from cell surface GPCRs to activation of perinuclear PLCε is not clear. Here we tested the role of G protein βγ subunits. Gβγ inhibition blocked ET-1-stimulated Golgi PI4P depletion in neonatal and adult ventricular myocytes. Blocking Gβγ at the Golgi inhibited ET-1-dependent PI4P depletion and nuclear PKD activation. Translocation of Gβγ to the Golgi stimulated perinuclear Golgi PI4P depletion and nuclear PKD activation. Finally, blocking Gβγ at the Golgi or PM blocked ET-1-dependent cardiomyocyte hypertrophy. These data indicate that Gβγ regulation of the perinuclear Golgi PI4P pathway and a separate pathway at the PM is required for ET-1-stimulated hypertrophy, and the efficacy of Gβγ inhibition in preventing heart failure maybe due in part to its blocking both these pathways.

Topics: Animals; Cardiomegaly; Cells, Cultured; Golgi Apparatus; GTP-Binding Protein beta Subunits; GTP-Binding Protein gamma Subunits; Hydrolysis; Myocytes, Cardiac; Phosphatidylinositol Phosphates; Protein Transport; Rats, Sprague-Dawley; Second Messenger Systems

Phospholipase Cε (PLCε) is a multifunctional enzyme implicated in cardiovascular, pancreatic, and inflammatory functions. Here we show that conditional deletion of PLCε in mouse cardiac myocytes protects from stress-induced pathological hypertrophy. PLCε small interfering RNA (siRNA) in ventricular myocytes decreases endothelin-1 (ET-1)-dependent elevation of nuclear calcium and activation of nuclear protein kinase D (PKD). PLCε scaffolded to muscle-specific A kinase-anchoring protein (mAKAP), along with PKCε and PKD, localizes these components at or near the nuclear envelope, and this complex is required for nuclear PKD activation. Phosphatidylinositol 4-phosphate (PI4P) is identified as a perinuclear substrate in the Golgi apparatus for mAKAP-scaffolded PLCε. We conclude that perinuclear PLCε, scaffolded to mAKAP in cardiac myocytes, responds to hypertrophic stimuli to generate diacylglycerol (DAG) from PI4P in the Golgi apparatus, in close proximity to the nuclear envelope, to regulate activation of nuclear PKD and hypertrophic signaling pathways.

Topics: Animals; Aorta; Cardiomegaly; Golgi Apparatus; GTP-Binding Protein alpha Subunits, Gq-G11; Heart; Heart Ventricles; Male; Mice; Myocytes, Cardiac; Nuclear Envelope; Phosphatidylinositol Phosphates; Phosphoinositide Phospholipase C; Rats; Signal Transduction