Mitochondrial fission is a highly regulated process mediated by a defined set of protein factors and is involved in the early stage of apoptosis. In mammals, at least two proteins, the dynamin-like protein DLP1/Drp1 and the mitochondrial outer membrane protein hFis1, participate in mitochondrial fission. The cytosolic domain of hFis1 contains six α-helices that form two tetratricopeptide repeat (TPR) motifs. Overexpression of hFis1 induces DLP1-mediated fragmentation of mitochondria, suggesting that hFis1 is a limiting factor in mitochondrial fission by recruiting cytosolic DLP1. In the present study, we identified two regions of hFis1 that are necessary for correct fission of mitochondria. We found that the TPR region of hFis1 participates in the interaction with DLP1 or DLP1-containing complex and that the first helix (α1) of hFis1 is required for mitochondrial fission presumably by regulating DLP1-hFis1 interaction. Misregulated interaction between DLP1 and hFis1 by α1 deletion induced mitochondrial swelling, in part by the mitochondrial permeability transition, but significantly delayed cell death. Our data suggest that hFis1 is a main regulator of mitochondrial fission, controlling the recruitment and assembly of DLP1 during both normal and apoptotic fission processes.

Our purpose was to evaluate hyporesponsivity to nitric oxide (NO)-induced dilation in small arterioles during nitrate tolerance. An Alza osmotic pump was implanted in the left flank of adult rats (n ϭ 56) for continuous administration of nitroglycerin (140 g/h) or vehicle (propylene glycol). On postoperative day 3, arcade (ϳ50-m diameter) and terminal (ϳ20 m) arterioles were observed in the cremaster preparation with in vivo video microscopy. Local vascular responses were obtained with micropipette-applied NO donors, with and without superoxide dismutase (SOD), Mn(III) tetrakis(4-benzoic acid) porphyrin chloride (MnTBAP), or losartan. On day 3, NO-mediated dilation was significantly attenuated in nitroglycerin-treated rats. Attenuation was greater in the terminal arterioles compared with the arcades. Control responses were restored by SOD, MnTBAP, or losartan, suggesting a role for elevated angiotensin II and reactive oxygen species (ROS) as mediators of the attenuated NO dilation (nitrate tolerance). Addition of losartan to the drinking water likewise prevented nitrate tolerance. In summary, terminal arterioles are affected by nitrates to a greater extent than the arcade arterioles that feed them, in a process dependent on angiotensin II and ROS. microvascular responses; steady vs. pulsatile flow; superoxide CLINICAL USE OF NITROGLYCERIN (NTG) to improve cardiac function in heart failure was proposed by Murrell in 1879 (28) and continues to be a treatment of choice to improve cardiac output through its peripheral dilatory effects, decreasing preload and afterload on the heart. Nitrate tolerance is a long-standing detrimental side effect of nitrate therapy involving vascular hyporesponsivity to continued nitrate treatment. In fact, cross-tolerance occurs, in which there is a failure of blood vessels to dilate to all nitric oxide (NO) donors; this is associated with an increase in constriction to many agents, including angiotensin II.In the systemic circulation, the mechanism of nitrate tolerance includes early activation of the renin-angiotensin system with elevation of vascular superoxide anion concentration (6,17,21,27). The cellular mechanism involves upregulation of angiotensin II and NADPH oxidase, with reduced expression and activity of superoxide dismutase (SOD) (26, 27). There is evidence that the diminished dilation to NO occurs via a process after cGMP synthesis (36). Recently, we showed (20) that cyclic nucleotide phosphodiesterase (PDE) 1A1 is upregulated in nitrate tolerance, suggesting that this cGMP-hydrolyzing PDE is involved in diminishing NO/cGMP-mediated vasodilation in tolerance. Thus our understanding of the cellular mechanism is improving. However, our understanding of nitrate tolerance in the whole animal is not yet complete.Most studies have examined nitrate tolerance with in vivo or in vitro models of the systemic circulation. Although the involvement of the peripheral circulation has been suggested and documented for many years, especially with regard to differences between vascular bed...

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Our purpose was to define arteriolar network hemodynamics during moderate increases in interstitial adenosine or nitric oxide in the hamster (n = 34, pentobarbital sodium 70 mg/kg) cheek pouch tissue. The network consists of a feed arteriole (approximately 12-microm diameter, approximately 800-microm length) with three to six branches. Observations of diameter, red blood cell flux, and velocity were obtained at the feed before the branch and within the branch. A comparison of baseline with suffused adenosine or sodium nitroprusside (SNP) 10(-9) to 10(-5) M showed the following. First, diameter change was heterogeneous by agonist, did not reflect the expected dilatory response, and was related to location within the network. With adenosine, upstream branch points constricted and those downstream dilated, even at 10(-5) M. With SNP, upstream branch points dilated, whereas those downstream constricted. Second, with adenosine, changes in diameter, flux, and velocity together resulted in no change in wall shear stress until 10(-5) M. Wall shear stress was not maintained at a constant level with Nomega-nitro-L-arginine (10(-5) M), suggesting a role for flow-dependent diameter changes with adenosine. With SNP, diameter change correlated with the baseline (before SNP) shear stress conditions.

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