An analysis of pH tolerance and substrate preference of isolated skeletal muscle mitochondria from Bufo marinus and Rana catesbeiana

1. The effects of varying pH and substrate on isolated skeletal muscle mitochondria from Bufo marinus and Rana catesbeiana were investigated. 2. For both species, VO2 max significantly decreased at all pH < 7.3 (P < 0.05), while maximum values were observed at a pH range of 7.3-7.6 with B. marinus maintaining a greater VO2 max than R. catesbeiana. 3. Respiratory control values (RCR) decreased significantly at all pH < 6.9 for both species (P < 0.05). 4. Isolated mitochondria from both species were maintained at pH = 7.2 and O2 consumption measured under five separate substrate conditions. 5. A rank preference was established based upon state 3 and RCR values. 6. Substrate preference was identical for both species and interspecific comparisons revealed differences in state 3 respiration and coupling.     

Dissecting cAMP binding domain A in the RIalpha subunit of cAMP-dependent protein kinase. Distinct subsites for recognition of cAMP and the catalytic subunit

The two gene-duplicated cAMP binding domains in the regulatory subunits of cAMP dependent protein kinase are each comprised of an A helix, an eight-stranded beta-barrel, and a B and C helix (1). The A domain is required for high affinity binding to C, while the B domain regulates access to the A domain. Using a combination of a yeast two-hybrid screen coupled with deletion analysis, cAMP binding domain A of RI was dissected into two structurally and functionally distinct subsites, one that binds cAMP and another that binds the C subunit. The minimum stable subdomain required for binding to C in the 1-3 micromolar range is composed of residues 94-169, while residues 236-244, mapped to the C helix of cAMP binding domain A, were defined as a second surface necessary for high affinity (5-10 nanomolar) binding to C. This portion of the C helix, due to its position directly between the two subsites, serves as a molecular switch for either a cAMP-bound conformation or a C-bound conformation and can thus modulate interactions of cAMP binding domain A with cAMP, with C, and with cAMP binding domain B.     

Transluminal balloon angioplasty, stents, and atherectomy

Arterial bypass grafting for occlusive disease is still considered the gold standard in the treatment of arterial occlusive disease. However, less invasive methods are available for select patients. Percutaneous balloon angioplasty has been shown to be effective in focal iliac artery stenosis, with patency rates of 50% to 90% at 5 years. Patency rates for femoropopliteal lesions are generally less than 50% at 2 years. Complications seen with balloon angioplasty often can be treated with arterial stents. Stents can be categorized into balloon-expandable, which provide a rigid scaffold to support the artery, self-expanding, which exert radial force to resist external compression, and thermal expanding stents, which allow for the use of a smaller introducer sheath. Another treatment option is atherectomy, in which the offending lesion is removed instead of fracturing and dilating the lumen. Although overall initial results of atherectomy have not been favorable, short lesions with eccentric atheroma or intimal hyperplasia, such as those seen in dialysis access fistulas or vein graft stenoses, may respond well.     

Aldosterone and dexamethasone stimulate calcineurin activity through a transcription-independent mechanism involving steroid receptor-associated heat shock proteins

1. PD 81,723 has been shown to enhance binding of adenosine to A1 receptors by stabilizing G protein-receptor coupling ('allosteric enhancement'). Evidence has been provided that in the perfused hearts and isolated atria PD 81,723 causes a sensitization to adenosine via this mechanism. 2. We have studied the effect of PD 81,723 in guinea-pig isolated atrial myocytes by use of whole-cell measurement of the muscarinic K+ current (I[K(ACh)]) activated by different Gi-coupled receptors (A1, M2, sphingolipid). PD 81,273 caused inhibition of I[K(ACh)] (IC50 approximately 5 microM) activated by either of the three receptors. Receptor-independent I[K(ACh)] in cells loaded with GTP-gamma-S and background I[K(ACh)], which contributes to the resting conductance of atrial myocytes, were equally sensitive to PD 81,723. At no combination of concentrations of adenosine and PD 81,723 could an enhancing effect be detected. 3. The compound was active from the outside only. Loading of the cells with PD 81,723 (50 microM) via the patch pipette did not affect either I[K(ACh)] or its sensitivity to adenosine. We suggest that PD 81,723 acts as an inhibitor of inward rectifying K+ channels; this is supported by the finding that ventricular I(K1), which shares a large degree of homology with the proteins (GIRK1/GIRK4) forming I[K(ACh)] but is not G protein-gated, was also blocked by this compound. 4. It is concluded that the functional effects of PD 81,723 described in the literature are not mediated by the A1 adenosine receptor-Gi-I[K(ACh)] pathway.     

Transducin-alpha C-terminal peptide binding site consists of C-D and E-F loops of rhodopsin

The binding of heterotrimeric GTP-binding proteins (G-proteins) to serpentine receptors involves several independent contacts. We have deduced the points of interaction between mutant bovine rhodopsins and alphat-(340-350), a peptide corresponding to the C terminus of the alpha subunit (alphat) of bovine retinal G-protein, transducin. Direct binding of alphat-(340-350) to rhodopsin stabilizes the activated metarhodopsin II state (M II), consequently uncoupling the rhodopsin-transducin interaction. This peptide action requires two segments on the cytoplasmic domain of rhodopsin: the Tyr136-Val137-Val138-Val139 sequence on the C-D loop and the Glu247-Lys248-Glu249-Val250-Thr251 sequence on the E-F loop. We propose that a tertiary interaction of these two loop regions forms a pocket for binding the alphat C terminus of the transducin during light transduction in vivo. In most G-proteins, the C termini of alpha subunits are important for interaction with receptors, and, in several serpentine receptors, regions similar to those in rhodopsin are essential for G-protein activation, indicating that the interaction described here may be a generally applicable mode of G-protein binding in signal transduction.     

Evaluation of the effectiveness of continuous renal replacement therapy in critical patients after cardiovascular surgery

The efficacy of continuous methods of renal substitute therapy (RST) in patients with multiple organ failure is assessed. The patients were divided in 2 groups administered different types of PST. Group 1 were 16 patients subjected to RST by peritoneal dialysis, in group 2 (n = 16) GP and/or GDP were used. Hemodynamics, hematological and biochemical values, and clearance of inflammation mediators were monitored and hemohydrobalance and complications of therapy assessed in the course of RST. Both RST methods proved to be highly effective. The possibility of differentiated use of peritoneal dialysis and GP/GDP permits an individual approach to treatment, and equally high efficacy of both methods solves the problem of treating total renal insufficiency in the majority of patients with multiple organ failure following cardiovascular surgery.