The relative changes in isometric force and work during fatigue and recovery in isolated toad sartorius muscle

Toad sartorius muscle was subjected to sinusoidal varying length changes at 2 Hz to measure work. Both isometric tetanic force and work per cycle were measured before, during, and after a 3-min fatigue. Both isometric tetanic force and positive work, the work done by the muscle during the shortening part of the cycle, rapidly decreased in parallel in the first 40 s of fatigue. Thereafter, force continued to decrease, but at a slower rate, to about 10% of prefatigue values, whereas positive work levelled off at about 30% of prefatigue values. Negative work, the work done on the muscle during the lengthening part of the cycle, increased during fatigue to the extent that net work became negative. This was due to a prolonged relaxation, which resulted in active force still being generated while the muscle was being stretched. Work and force recovered at about the same rate. Isometric force measurements alone do not give any clear indication that net work will be negative under a particular set of experimental conditions.     

Clinical and Molecular Characterization of Patients with Distal 11q Deletions

Jacobsen syndrome is caused by segmental aneusomy for the distal end of the long arm of chromosome 11. Typical features include mild to moderate psychomotor retardation, trigonocephaly, facial dysmorphism, cardiac defects, and thrombocytopenia, though none of these features are invariably present. To define the critical regions responsible for these abnormalities, we studied 17 individuals with de novo terminal deletions of 11q. The patients were characterized in a loss-of-heterozygosity analysis using polymorphic dinucleotide repeats. The breakpoints in the complete two-generation families were localized with an average resolution of 3.9 cM. Eight patients with the largest deletions extending from 11q23.3 to 11qter have breakpoints, between D11S924 and D11S1341. This cytogenetic region accounts for the majority of 11q⁻ patients and may be related to the FRA11B fragile site in 11q23.3. One patient with a small terminal deletion distal to D11S1351 had facial dysmorphism, cardiac defects, and thrombocytopenia, suggesting that the genes responsible for these features may lie distal to D11S1351. Twelve of 15 patients with deletion breakpoints as far distal as D11S1345 had trigonocephaly, while patients with deletions distal to D11S912 did not, suggesting that, if hemizygosity for a single gene is responsible for this dysmorphic feature, the gene may lie distal to D11S1345 and proximal to D11S912.     

Magnesium-induced inner membrane aggregation in heart mitochondria: prevention and reversal by carboxyatractyloside and bongkrekic acid

Mg(2+) at an optimal concentration of 2mM (ph 6.5) induces large increases (up to 30 percent) in the optical density of bovine heart mitochondria incubated under conditions of low ionic strength (< approx. 0.01). The increases are associated with aggregation (sticking together) of the inner membranes and are little affected by changes in the energy status of the mitochondria. Virtually all of a number of other polyvalent cations tested and Ag(+) induce increases in mitochondrial optical density similar to those induced by Mg(2+), their approximate order of concentration effectiveness in respect to Mg(2+) being: La(3+) > Pb(2+) = Cu(2+) > Cd(2+) > Zn(2+) > Ag(+) > Mn(2+) > Ca(2+) > Mg(2+). With the exception of Mg(2+), all of these cations appear to induce swelling of the mitochondria concomitant with inner membrane aggregation. The inhibitors of the adenine nucleotide transport reaction carboxyatratyloside and bongkrekic acid are capable of preventing and reversing Mg(2+)-induced aggregation at the same low concentration required for complete inhibition of phosphorylating respiration, suggesting that they inhibit the aggregation by binding to the adenine nucleotide carrier. The findings are interpreted to indicate (a) that the inner mitochondrial membrane is normally prevented from aggregating by virtue of its net negative outer surface change, (b) that the cations induce the membrane to aggregate by binding at its outer surface, decreasing the net negative charge, and (c) that carboxyatractyloside and bongkrekic acid inhibit the aggregation by binding to the outer surface of the membrane, increasing the net negative charge.     

Scaling-up the delivery of dog vaccination campaigns against rabies in Tanzania

An increasing number of countries are committing to meet the global target to eliminate human deaths from dog-mediated rabies by 2030. Mass dog vaccination is central to this strategy. To interrupt rabies transmission from dogs to humans, the World Health Organization recommends that vaccination campaigns should be carried out every year in all dog-owning communities vaccinating 70% of their susceptible dogs. Monitoring and evaluation of dog vaccination campaigns are needed to measure progress towards elimination. In this study, we measured the delivery performance of large-scale vaccination campaigns implemented in 25 districts in south-east Tanzania from 2010 until 2017. We used regression modelling to infer the factors associated with, and potentially influencing the successful delivery of vaccination campaigns. During 2010–2017, five rounds of vaccination campaigns were carried out, vaccinating in total 349,513 dogs in 2,066 administrative vaccination units (rural villages or urban wards). Progressively more dogs were vaccinated over the successive campaigns. The campaigns did not reach all vaccination units each year, with only 16–28% of districts achieving 100% campaign completeness (where all units were vaccinated). During 2013–2017 when vaccination coverage was monitored, approximately 20% of vaccination units achieved the recommended 70% coverage, with average coverage around 50%. Campaigns were also not completed at annual intervals, with the longest interval between campaigns being 27 months. Our analysis revealed that districts with higher budgets generally achieved higher completeness, with a twofold difference in district budget increasing the odds of a vaccination unit being reached by a campaign by slightly more than twofold (OR: 2.29; 95% CI: 1.69–3.09). However, higher budgets did not necessarily result in higher coverage within vaccination units that were reached. We recommend national programs regularly monitor and evaluate the performance of their vaccination campaigns, so as to identify factors hindering their effective delivery and to guide remedial action.     

Molecular localization of the inhibitory arachidonic acid binding site to the pore of hIK1

We previously demonstrated that the endogenously expressed human intermediate conductance, Ca(2+)-activated K(+) channel (hIK1) was inhibited by arachidonic acid (AA) (Devor, D. C., and Frizzell, R. A. (1998) Am. J. Physiol. 274, C138-C148). Here we demonstrate, using the excised, inside-out patch-clamp technique, that hIK1, heterologously expressed in HEK293 cells, is inhibited 82 +/- 2% (n = 16) with 3 microm AA, being half-maximally inhibited (IC(50)) at 1.4 +/- 0.7 microm. In contrast, AA does not inhibit the Ca(2+)-dependent, small conductance K(+) channel, rSK2, another member of the KCNN gene family. Therefore, we utilized chimeric hIK1/rSK2 channels to define the AA binding domain on hIK1 to the S5-Pore-S6 region of the channel. Subsequent site-directed mutagenesis revealed that mutation of Thr(250) to Ser (T250S) resulted in a channel with limited sensitivity to block by AA (8 +/- 2%, n = 8), demonstrating that Thr(250) is a key molecular determinant for the inhibition of hIK1 by AA. Likewise, when Val(275) in S6 was mutated to Ala (V275A) AA inhibited only 43 +/- 11% (n = 9) of current flow. The double mutation T250S/V275A eliminated the AA sensitivity of hIK1. Introducing the complimentary single amino acid substitutions into rSK2 (S359T and A384V) conferred partial AA sensitivity to rSK2, 21 +/- 3% and 31 +/- 3%, respectively. Further, introducing the double mutation S359T/A384V into rSK2 resulted in a 63 +/- 8% (n = 9) inhibition by AA, thereby demonstrating the ability to introduce this inhibitory AA binding site into another member of the KCNN gene family. These results demonstrate that AA interacts with the pore-lining amino acids, Thr(250) and Val(275) in hIK1, conferring inhibition of hIK1 by AA and that AA and clotrimazole share similar, if not identical, molecular sites of interaction.