Nitric oxide was found to trigger mitochondrial biogenesis in cells as diverse as brown adipocytes and 3T3-L1, U937, and HeLa cells. This effect of nitric oxide was dependent on guanosine 3′,5′-monophosphate (cGMP) and was mediated by the induction of peroxisome proliferator–activated receptor γ coactivator 1α, a master regulator of mitochondrial biogenesis. Moreover, the mitochondrial biogenesis induced by exposure to cold was markedly reduced in brown adipose tissue of endothelial nitric oxide synthase null-mutant (eNOS ^–/– ) mice, which had a reduced metabolic rate and accelerated weight gain as compared to wild-type mice. Thus, a nitric oxide–cGMP–dependent pathway controls mitochondrial biogenesis and body energy balance.

We have earlier shown that microglia, the immune cells of the CNS, release microparticles from cell plasma membrane after ATP stimulation. These vesicles contain and release IL‐1β, a crucial cytokine in CNS inflammatory events. In this study, we show that microparticles are also released by astrocytes and we get insights into the mechanism of their shedding. We show that, on activation of the ATP receptor P2X7, microparticle shedding is associated with rapid activation of acid sphingomyelinase, which moves to plasma membrane outer leaflet. ATP‐induced shedding and IL‐1β release are markedly reduced by the inhibition of acid sphingomyelinase, and completely blocked in glial cultures from acid sphingomyelinase knockout mice. We also show that p38 MAPK cascade is relevant for the whole process, as specific kinase inhibitors strongly reduce acid sphingomyelinase activation, microparticle shedding and IL‐1β release. Our results represent the first demonstration that activation of acid sphingomyelinase is necessary and sufficient for microparticle release from glial cells and define key molecular effectors of microparticle formation and IL‐1β release, thus, opening new strategies for the treatment of neuroinflammatory diseases.

The endoplasmic reticulum (ER) can transform from a network of branching tubules into stacked membrane arrays (termed organized smooth ER [OSER]) in response to elevated levels of specific resident proteins, such as cytochrome b(5). Here, we have tagged OSER-inducing proteins with green fluorescent protein (GFP) to study OSER biogenesis and dynamics in living cells. Overexpression of these proteins induced formation of karmellae, whorls, and crystalloid OSER structures. Photobleaching experiments revealed that OSER-inducing proteins were highly mobile within OSER structures and could exchange between OSER structures and surrounding reticular ER. This indicated that binding interactions between proteins on apposing stacked membranes of OSER structures were not of high affinity. Addition of GFP, which undergoes low affinity, antiparallel dimerization, to the cytoplasmic domains of non–OSER-inducing resident ER proteins was sufficient to induce OSER structures when overexpressed, but addition of a nondimerizing GFP variant was not. These results point to a molecular mechanism for OSER biogenesis that involves weak homotypic interactions between cytoplasmic domains of proteins. This mechanism may underlie the formation of other stacked membrane structures within cells.

We recently found that long-term exposure to nitric oxide (NO) triggers mitochondrial biogenesis in mammalian cells and tissues by activation of guanylate cyclase and generation of cGMP. Here, we report that the NO/cGMP-dependent mitochondrial biogenesis is associated with enhanced coupled respiration and content of ATP in U937, L6, and PC12 cells. The observed increase in ATP content depended entirely on oxidative phosphorylation, because ATP formation by glycolysis was unchanged. Brain, kidney, liver, heart, and gastrocnemius muscle from endothelial NO synthase null mutant mice displayed markedly reduced mitochondrial content associated with significantly lower oxygen consumption and ATP content. In these tissues, ultrastructural analyses revealed significantly smaller mitochondria. Furthermore, a significant reduction in the number of mitochondria was observed in the subsarcolemmal region of the gastrocnemius muscle. We conclude that NO/cGMP stimulates mitochondrial biogenesis, both in vitro and in vivo , and that this stimulation is associated with increased mitochondrial function, resulting in enhanced formation of ATP.