Intercellular calcium wave propagation initiated by mechanical stress is a phenomenon found in nearly all cell types. The waves utilize two pathways: transfer of InsP3 directly from cell to cell through gap junction channels and release of ATP onto extracellular purinergic receptors. The conduit for ATP has remained elusive and both a vesicular and a channel mediated release have been considered. Here, we describe the properties of single pannexin 1 channels. They have a wide expression spectrum, they are of large conductance and permeant for ATP, and they are mechanosensitive. Hence, pannexins are candidates for the release of ATP to the extracellular space upon mechanical stress.

ATP is a widely used extracellular signaling molecule. The mechanism of ATP release from cells is presently unresolved and may be either vesicular or channel-mediated. Erythrocytes release ATP in response to low oxygen or to shear stress. In the absence of vesicles, the release has to be through channels. Erythrocytes do not form gap junctions. Yet, here we show with immunohistochemical and electrophysiological data that erythrocytes express the gap junction protein pannexin 1. This protein, in addition to forming gap junction channels in paired oocytes, can also form a mechanosensitive and ATP-permeable channel in the nonjunctional plasma membrane. Consistent with a role of pannexin 1 as an ATP release channel, ATP release by erythrocytes was attenuated by the gap junction blocker carbenoxolone. Furthermore, under conditions of ATP release, erythrocytes took up fluorescent tracer molecules permeant to gap junction channels.ATP release ͉ gap junction ͉ hemichannel ͉ dye uptake P annexins represent a recently discovered second family of gap junction proteins in vertebrates (1). Pannexins have no sequence homology with the well known connexin family of vertebrate gap junction proteins. Instead, they are related to innexins, which were originally considered to be exclusively invertebrate gap junction proteins. The functional role of pannexins is unknown. The existence of connexin-specific diseases, despite an overlap of connexin and pannexin expression, suggests a functional role of pannexins that is distinct from that of connexins. Pannexin 1, in addition to forming gap junctions in paired oocytes, also forms nonjunctional membrane channels that provide a passageway from the cytoplasm to the extracellular space for molecules in the size range of second messengers (2, 3). It can be hypothesized that the physiological role of pannexin 1 is formation of a nonjunctional membrane channel.Although the role of ATP as an extracellular signaling molecule is well recognized (4), the release mechanism for ATP from cells to the extracellular space has remained enigmatic. Two general release modes have been proposed: (i) vesicular release akin to the exocytotic release of transmitters and (ii) channel-mediated release. Although vesicular ATP release is well documented (5, 6), it cannot account for all of the ATP release phenomena. In particular, ATP is released from erythrocytes, which, under physiological conditions, are vesicle-free (7). Various channels have been implicated in the process, including CFTR (cystic fibrosis transmembrane conductance regulator), connexin 43 (Cx43) hemichannels, a volumeregulated channel (VRAC), and the purinergic receptor P2X7 (5,6,(8)(9)(10)(11). However, the evidence for their involvement falls short because of questionable specificity of the pharmacological blockers used to determine channel identity.Mechanical stress is a prime stimulus for ATP release in many cell types, including erythrocytes (12). An efficient release mechanism thus may involve a channel that is both mechanosensitive and...

Insulin activates insulin receptors (InsRs) in the hypothalamus to signal satiety after a meal. However, the rising incidence of obesity, which results in chronically elevated insulin levels, implies that insulin may also act in brain centres that regulate motivation and reward. We report here that insulin can amplify action potential-dependent dopamine (DA) release in the nucleus accumbens (NAc) and caudate–putamen through an indirect mechanism that involves striatal cholinergic interneurons that express InsRs. Furthermore, two different chronic diet manipulations in rats, food restriction (FR) and an obesogenic (OB) diet, oppositely alter the sensitivity of striatal DA release to insulin, with enhanced responsiveness in FR, but loss of responsiveness in OB. Behavioural studies show that intact insulin levels in the NAc shell are necessary for acquisition of preference for the flavour of a paired glucose solution. Together, these data imply that striatal insulin signalling enhances DA release to influence food choices.

Key Points• Risk stratification treatment of t(8;21) acute myeloid leukemia may decrease relapse and improve longterm survival.• Allo-HSCT benefited high-risk patients, but impaired the survival of low-risk patients.We aimed to improve the outcome of t(8;21) acute myeloid leukemia (AML) in the first complete remission (CR1) by applying risk-directed therapy based on minimal residual disease (MRD) determined by RUNX1/RUNX1T1 transcript levels. Risk-directed therapy included recommending allogeneic hematopoietic stem cell transplantation (allo-HSCT) for high-risk patients and chemotherapy/autologous-HSCT (auto-HSCT) for low-risk patients. Among 116 eligible patients, MRD status after the second consolidation rather than induction or first consolidation could discriminate high-risk relapse patients (P 5 .001).Allo-HSCT could reduce relapse and improve survival compared with chemotherapy for high-risk patients (cumulative incidence of relapse [CIR]: 22.1% vs 78.9%, P < .0001; disease-free survival [DFS]: 61.7% vs 19.6%, P 5 .001), whereas chemotherapy/auto-HSCT achieved a low relapse rate (5.3%) and high DFS (94.7%) for low-risk patients. Multivariate analysis revealed that MRD status and treatment choice were independent prognostic factors for relapse, DFS, and OS. We concluded that MRD status after the second consolidation may be the best timing for treatment choice. MRD-directed risk stratification treatment may improve the outcome of t(8;21) AML in CR1. This trial was registered at http://www.chictr.org as #ChiCTR- OCH-12002406. (Blood. 2013;121(20):4056-4062)

Mitochondrial dysfunction is a potential causal factor in Parkinson's disease. We show here that acute exposure to the mitochondrial complex I inhibitor rotenone (30 -100 nM; 30 min) causes concentration-dependent suppression of single-pulse evoked dopamine (DA) release monitored in real time with carbon-fiber microelectrodes in guinea pig striatal slices, with no effect on DA content. Suppression of DA release was prevented by the sulfonylurea glibenclamide, implicating ATP-sensitive K ϩ (K ATP ) channels; however, tissue ATP was unaltered. Because K ATP channels can be activated by hydrogen peroxide (H 2 O 2 ), as well as by low ATP, we examined the involvement of rotenone-enhanced H 2 O 2 generation. Confirming an essential role for H 2 O 2 , the inhibition of DA release by rotenone was prevented by catalase, a peroxide-scavenging enzyme. Striatal H 2 O 2 generation during rotenone exposure was examined in individual medium spiny neurons using fluorescence imaging with dichlorofluorescein (DCF). An increase in intracellular H 2 O 2 levels followed a similar time course to that of DA release suppression and was accompanied by cell membrane depolarization, decreased input resistance, and increased excitability. Extracellular catalase markedly attenuated the increase in DCF fluorescence and prevented rotenone-induced effects on membrane properties; membrane changes were also largely prevented by flufenamic acid, a blocker of transient receptor potential (TRP) channels. Thus, partial mitochondrial inhibition can cause functional DA denervation via H 2 O 2 and K ATP channels, without DA or ATP depletion. Furthermore, amplified H 2 O 2 levels and TRP channel activation in striatal spiny neurons indicate potential sources of damage in these cells. Overall, these novel factors could contribute to parkinsonian motor deficits and neuronal degeneration caused by mitochondrial dysfunction.