Phosphodiesterase 2A as a therapeutic target to restore cardiac neurotransmission during sympathetic hyperactivity
Kun Liu, Dan Li, Guoliang Hao, David McCaffary, Oliver Neely, Lavinia Woodward, Demetris Ioannides, Chieh-Ju Lu, Marcella Brescia, Manuela Zaccolo, Harikrishna Tandri, Olujimi A. Ajijola, Jeffrey L. Ardell, Kalyanam Shivkumar, David J. Paterson
Kun Liu, Dan Li, Guoliang Hao, David McCaffary, Oliver Neely, Lavinia Woodward, Demetris Ioannides, Chieh-Ju Lu, Marcella Brescia, Manuela Zaccolo, Harikrishna Tandri, Olujimi A. Ajijola, Jeffrey L. Ardell, Kalyanam Shivkumar, David J. Paterson
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Research Article Cardiology Neuroscience

Phosphodiesterase 2A as a therapeutic target to restore cardiac neurotransmission during sympathetic hyperactivity

Abstract

Elevated levels of brain natriuretic peptide (BNP) are regarded as an early compensatory response to cardiac myocyte hypertrophy, although exogenously administered BNP shows poor clinical efficacy in heart failure and hypertension. We tested whether phosphodiesterase 2A (PDE2A), which regulates the action of BNP-activated cyclic guanosine monophosphate (cGMP), was directly involved in modulating Ca2+ handling from stellate ganglia (SG) neurons and cardiac norepinephrine (NE) release in rats and humans with an enhanced sympathetic phenotype. SG were also isolated from patients with sympathetic hyperactivity and healthy donor patients. PDE2A activity of the SG was greater in both spontaneously hypertensive rats (SHRs) and patients compared with their respective controls, whereas PDE2A mRNA was only high in SHR SG. BNP significantly reduced the magnitude of the calcium transients and ICaN in normal Wistar Kyoto (WKY) SG neurons, but not in the SHRs. cGMP levels stimulated by BNP were also attenuated in SHR SG neurons. Overexpression of PDE2A in WKY neurons recapitulated the calcium phenotype seen in SHR neurons. Functionally, BNP significantly reduced [3H]-NE release in the WKY rats, but not in the SHRs. Blockade of overexpressed PDE2A with Bay 60-7550 or overexpression of catalytically inactive PDE2A reestablished the modulatory action of BNP in SHR SG neurons. This suggests that PDE2A may be a key target in modulating the action of BNP to reduce sympathetic hyperactivity.

Authors

Kun Liu, Dan Li, Guoliang Hao, David McCaffary, Oliver Neely, Lavinia Woodward, Demetris Ioannides, Chieh-Ju Lu, Marcella Brescia, Manuela Zaccolo, Harikrishna Tandri, Olujimi A. Ajijola, Jeffrey L. Ardell, Kalyanam Shivkumar, David J. Paterson

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Figure 4

Measurement of calcium current and intracellular free calcium transients in stellate ganglia neurons in the presence of BNP and phosphodiesterase 2A inhibitor.

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Measurement of calcium current and intracellular free calcium transients...
(A) Fluorescence images of a cultured cardiac sympathetic neurons derived from a 4-week-old Wistar Kyoto (WKY) rat stellate ganglion that was stained for the catecholamine neuronal marker tyrosine hydroxylase (TH, red), phosphodiesterase 2A (PDE2A) antibody (green), costained with the nuclear marker DAPI (blue), and overlay. Scale bar: 20 μm. (B) Representative whole-cell calcium current traces (upper graphs) obtained before and after exposure to 100 nmol/l brain natriuretic peptide (BNP) (left, middle) or 100 nmol/l BNP with 1 μmol/l Bay 60-7550 (right) from 4-week-old WKY rats and spontaneously hypertensive rats (SHRs). Currents were evoked by test pulses of −10 mV from a holding potential of –90 mV. Mean current density–voltage relationships (lower graphs) in the presence and absence of 100 nmol/l BNP (left, middle) or 100 nmol/l BNP with 1 μmol/l Bay 60-7550 (right) from 4-week-old WKY rats and SHRs. *P < 0.05 by paired t test. Vm, membrane voltage in mV; ICa indicates calcium current; pA, picoampere; pF, picofarad. (C) Pharmacological protocols (upper) and raw data trace (lower) recording intracellular calcium transient ([Ca2+]i) in a single cardiac sympathetic neuron from 4-week-old WKY rats and SHRs. A neuron loaded with fura-2 acetoxymethyl ester (Fura-2/AM, 2 μmol/l) was stimulated by 50 mmol/l KCl for 30 seconds to depolarize the neuron and evoke voltage-gated Ca2+ entry. The size of the first (S1) and second (S2) KCl stimulation was compared. (D) Group data showing KCl-evoked peak [Ca2+]i changes expressed as a ratio (%) of S2 compared with S1 in response to 100 or 250 nmol/l BNP with or without PDE2 inhibitor Bay 60-7550 (1 μmol/l) in the WKY rats (upper) and SHRs (lower). *P < 0.05, **P < 0.01, ***P < 0.001, compared with control; †P < 0.05, †††P < 0.001; one-way ANOVA. n indicates the number of neurons.