Pulmonary arterial hypertension is a devastating disease with high mortality. Familial cases of pulmonary arterial hypertension are usually characterized by autosomal dominant transmission with reduced penetrance, and some familial cases have unknown genetic causes. We studied a family in which multiple members had pulmonary arterial hypertension without identifiable mutations in any of the genes known to be associated with the disease, including BMPR2, ALK1, ENG, SMAD9, and CAV1. Three family members were studied with whole-exome sequencing. Additional patients with familial or idiopathic pulmonary arterial hypertension were screened for the mutations in the gene that was identified on whole-exome sequencing. All variants were expressed in COS-7 cells, and channel function was studied by means of patch-clamp analysis. We identified a novel heterozygous missense variant c.608 G→A (G203D) in KCNK3 (the gene encoding potassium channel subfamily K, member 3) as a disease-causing candidate gene in the family. Five additional heterozygous missense variants in KCNK3 were independently identified in 92 unrelated patients with familial pulmonary arterial hypertension and 230 patients with idiopathic pulmonary arterial hypertension. We used in silico bioinformatic tools to predict that all six novel variants would be damaging. Electrophysiological studies of the channel indicated that all these missense mutations resulted in loss of function, and the reduction in the potassium-channel current was remedied by the application of the phospholipase inhibitor ONO-RS-082. Our study identified the association of a novel gene, KCNK3, with familial and idiopathic pulmonary arterial hypertension. Mutations in this gene produced reduced potassium-channel current, which was successfully remedied by pharmacologic manipulation. (Funded by the National Institutes of Health.)

Morphine is one of the most prescribed and effective drugs used for the treatment of acute and chronic pain conditions. In addition to its central effects, morphine can also produce peripheral analgesia. However, the mechanisms underlying this peripheral action of morphine have not yet been fully elucidated. Here, we show that the peripheral antinociceptive effect of morphine is lost in neuronal nitric-oxide synthase null mice and that morphine induces the production of nitric oxide in primary nociceptive neurons. The activation of the nitric-oxide pathway by morphine was dependent on an initial stimulation of PI3Kγ/AKT protein kinase B (AKT) and culminated in increased activation of K _ATP channels. In the latter, this intracellular signaling pathway might cause a hyperpolarization of nociceptive neurons, and it is fundamental for the direct blockade of inflammatory pain by morphine. This understanding offers new targets for analgesic drug development.

Overwhelming evidence supports the importance of the sympathetic nervous system in heart failure. In contrast, much less is known about the role of failing cholinergic neurotransmission in cardiac disease. By using a unique genetically modified mouse line with reduced expression of the vesicular acetylcholine transporter (VAChT) and consequently decreased release of acetylcholine, we investigated the consequences of altered cholinergic tone for cardiac function. M-mode echocardiography, hemodynamic experiments, analysis of isolated perfused hearts, and measurements of cardiomyocyte contraction indicated that VAChT mutant mice have decreased left ventricle function associated with altered calcium handling. Gene expression was analyzed by quantitative reverse transcriptase PCR and Western blotting, and the results indicated that VAChT mutant mice have profound cardiac remodeling and reactivation of the fetal gene program. This phenotype was attributable to reduced cholinergic tone, since administration of the cholinesterase inhibitor pyridostigmine for 2 weeks reversed the cardiac phenotype in mutant mice. Our findings provide direct evidence that decreased cholinergic neurotransmission and underlying autonomic imbalance cause plastic alterations that contribute to heart dysfunction.

Infection with Trypanosoma cruzi induces inflammation, which limits parasite proliferation but may result in chagasic heart disease. Suppressor of cytokine signaling 2 (SOCS2) is a regulator of immune responses and may therefore participate in the pathogenesis of T. cruzi infection. SOCS2 is expressed during T. cruzi infection, and its expression is partially reduced in infected 5-lipoxygenase-deficient [knockout (KO)] mice. In SOCS2 KO mice, there was a reduction in both parasitemia and the expression of interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), IL-6, IL-10, SOCS1, and SOCS3 in the spleen. Expression of IFN-γ, TNF-α, SOCS1, and SOCS3 was also reduced in the hearts of infected SOCS2 KO mice. There was an increase in the generation and expansion of T regulatory (Treg) cells and a decrease in the number of memory cells in T. cruzi-infected SOCS2 KO mice. Levels of lipoxinA(4) (LXA(4)) increased in these mice. Echocardiography studies demonstrated an impairment of cardiac function in T. cruzi-infected SOCS2 KO mice. There were also changes in calcium handling and in action potential waveforms, and reduced outward potassium currents in isolated cardiac myocytes. Our data suggest that reductions of inflammation and parasitemia in infected SOCS2-deficient mice may be secondary to the increases in Treg cells and LXA(4) levels. This occurs at the cost of greater infection-associated heart dysfunction, highlighting the relevance of balanced inflammatory and immune responses in preventing severe T. cruzi-induced disease.

Background: Heterozygous loss of function mutations in the KCNK 3 gene cause hereditary pulmonary arterial hypertension ( PAH ). KCNK 3 encodes an acid‐sensitive potassium channel, which contributes to the resting potential of human pulmonary artery smooth muscle cells. KCNK 3 is widely expressed in the body, and dimerizes with other KCNK 3 subunits, or the closely related, acid‐sensitive KCNK 9 channel. Methods and Results: We engineered homomeric and heterodimeric mutant and nonmutant KCNK 3 channels associated with PAH . Using whole‐cell patch‐clamp electrophysiology in human pulmonary artery smooth muscle and COS 7 cell lines, we determined that homomeric and heterodimeric mutant channels in heterozygous KCNK 3 conditions lead to mutation‐specific severity of channel dysfunction. Both wildtype and mutant KCNK 3 channels were activated by ONO ‐ RS ‐082 (10 μmol/L), causing cell hyperpolarization. We observed robust gene expression of KCNK 3 in healthy and familial PAH patient lungs, but no quantifiable expression of KCNK 9 , and demonstrated in functional studies that KCNK 9 minimizes the impact of select KCNK 3 mutations when the 2 channel subunits co‐assemble. Conclusions: Heterozygous KCNK 3 mutations in PAH lead to variable loss of channel function via distinct mechanisms. Homomeric and heterodimeric mutant KCNK 3 channels represent novel therapeutic substrates in PAH . Pharmacological and pH ‐dependent activation of wildtype and mutant KCNK 3 channels in pulmonary artery smooth muscle cells leads to membrane hyperpolarization. Co‐assembly of KCNK 3 with KCNK 9 subunits may provide protection against KCNK 3 loss of function in tissues where both KCNK 9 and KCNK 3 are expressed, contributing to the lung‐specific phenotype observed clinically in patients with PAH because of KCNK 3 mutations.