Non-Invasive In Vivo Imaging of Calcium Signaling in Mice

Rapid and transient elevations of Ca2+ within cellular microdomains play a critical role in the regulation of many signal transduction pathways. Described here is a genetic approach for non-invasive detection of localized Ca2+ concentration ([Ca2+]) rises in live animals using bioluminescence imaging (BLI). Transgenic mice conditionally expressing the Ca2+-sensitive bioluminescent reporter GFP-aequorin targeted to the mitochondrial matrix were studied in several experimental paradigms. Rapid [Ca2+] rises inside the mitochondrial matrix could be readily detected during single-twitch muscle contractions. Whole body patterns of [Ca2+] were monitored in freely moving mice and during epileptic seizures. Furthermore, variations in mitochondrial [Ca2+] correlated to behavioral components of the sleep/wake cycle were observed during prolonged whole body recordings of newborn mice. This non-invasive imaging technique opens new avenues for the analysis of Ca2+ signaling whenever whole body information in freely moving animals is desired, in particular during behavioral and developmental studies

Heterogeneous natural selection on oxidative phosphorylation genes among fishes with extreme high and low aerobic performance

Background: Oxidative phosphorylation (OXPHOS) is the primary source of ATP in eukaryotes and serves as a mechanistic link between variation in genotypes and energetic phenotypes. While several physiological and anatomical factors may lead to increased aerobic capacity, variation in OXPHOS proteins may influence OXPHOS efficiency and facilitate adaptation in organisms with varied energy demands. Although there is evidence that natural selection acts on OXPHOS genes, the focus has been on detection of directional (positive) selection on specific phylogenetic branches where traits that increase energetic demands appear to have evolved. We examined patterns of selection in a broader evolutionary context, i.e., on multiple lineages of fishes with extreme high and low aerobic performance. Results: We found that patterns of natural selection on mitochondrial OXPHOS genes are complex among fishes with different swimming performance. Positive selection is not consistently associated with high performance taxa and appears to be strongest on lineages containing low performance taxa. In contrast, within high performance lineages, purifying (negative) selection appears to predominate. Conclusions: We provide evidence that selection on OXPHOS varies in both form and intensity within and among lineages through evolutionary time. These results provide evidence for fluctuating selection on OXPHOS associated with divergence in aerobic performance. However, in contrast to previous studies, positive selection was strongest on low performance taxa suggesting that adaptation of OXPHOS involves many factors beyond enhancing ATP production in high performance taxa. The broader pattern indicates a complex interplay between organismal adaptations, ATP demand, and OXPHOS function.This work was supported by NSF award DEB-0732988 (to REB).Ye

Subcutaneous sumatriptan delivery devices: comparative ease of use and preference among migraineurs

Anthony D Andre,1 Elimor Brand-Schieber,2 Margarita Ramirez,1 Sagar Munjal,2 Rajesh Kumar2 1Interface Analysis Associates, Saraftoga, CA, 2Dr Reddy’s Laboratories Inc., Princeton, NJ, USA Background: Several sumatriptan subcutaneous autoinjector devices for acute treatment of migraine patients are available, each device differs with respect to design and features. Determining device preference and ease of use is important because patients experiencing a migraine attack are often functionally impaired. Objective: The objective of this human factors study was to compare migraine patients’ device use performance and preferences for three sumatriptan subcutaneous autoinjectors: a disposable two-step device (Zembrace® SymTouch®), a disposable three-step device (Sumavel® DosePro®), and a multistep reloadable device (Imitrex® STATdose®), using simulated injections. Methods: Each study subject performed two unaided simulated injections with each of three different drug delivery devices, which were presented in counterbalanced order. The participants were then asked to rate the three devices on various subjective measures. The primary end point was overall device preference using a visual analog scale. Results: A total of 54 subjects participated and each subject performed two simulated injections with each of the three devices. Most subjects preferred the two-step device (88.9%) to the three-step (13.0%) and the reloadable (1.9%). The two-step device had higher mean overall preference ratings (F (2, 159)=56.6, P<0.01) and higher ratings for ease of use, intuitiveness, convenience, portability, and control. The two-step device had a first injection full-dose delivery success rate of 44.4%, higher than both the reloadable (24.1%) and the three-step (3.7%) devices. The number of errors with the two-step device (n=3) was ~90% lower than the three-step (n=49) and reloadable (n=44) devices. Conclusion: In this human factors study, 54 migraineurs used simulated injections to compare three sumatriptan subcutaneous delivery devices. Zembrace SymTouch, a two-step device, was most preferred compared with Sumavel DosePro and Imitrex STATdose. It also ranked highest for ease of use and various other measures. In this study, migraine patients preferred the autoinjector that they rated as simpler and more intuitive. Keywords: human factors, autoinjector, sumatriptan, migraine, preferenc

Effect of renal impairment on the pharmacokinetics of levomilnacipran following a single oral dose of levomilnacipran extended-release capsule in humans

Laishun Chen, William M Greenberg, Elimor Brand-Schieber, Julie Wangsa, Antonia Periclou, Parviz Ghahramani Forest Research Institute, a subsidiary of Actavis Inc., Jersey City, NJ, USA Purpose: Levomilnacipran extended-release (ER) is indicated for treatment of major depressive disorder in adults. We evaluated the pharmacokinetic and safety profile of levomilnacipran ER in individuals with impaired renal function. Methods: A total of 32 individuals participated in four groups (eight in each group) with normal, mild, moderately, or severely impaired renal function. Each participant received one dose of levomilnacipran ER 40 mg. Blood and urine were assayed using liquid chromatography/tandem mass spectrometry. Results between normal and renally impaired groups were compared using analysis of variance. Safety measures included adverse events, laboratory evaluations, vital signs, suicidality, and electrocardiograms. Results: Following administration of levomilnacipran, mean (standard deviation) maximum plasma concentration in participants with normal renal function, and mild, moderate, or severe renal impairment was 83.9 (21.0), 81.8 (23.4), 98.7 (18.1), and 122.1 (35.1) (ng/mL), respectively; area under the curve from time zero to infinity was 2,101.0 (516.9), 2,587.8 (649.9), 4,016.4 (995.4), and 5,900.8 (1,799.3) (h·ng/mL), respectively; terminal elimination half-life was 13.5 (2.8), 17.3 (3.5), 19.1 (4.6), and 27.7 (7.4) (hours), respectively; and renal clearance was 175.9 mL/min, 114.7 mL/min, 69.9 mL/min, and 28.6 mL/min, respectively. Levomilnacipran ER was generally well tolerated with no safety issues of concern identified. Conclusion: Renal impairment was associated with increased plasma levels of levomilnacipran and prolonged half-life. No dose adjustment is required for individuals with mild renal impairment; the recommended maximum daily maintenance dose of levomilnacipran ER should not exceed 80 mg for individuals with moderate renal impairment and 40 mg for individuals with severe renal impairment. Keywords: antidepressant, F2695, SNRI, pharmacokinetics, renal function, major depressive disorde

Human factors validation study of 3 mg sumatriptan autoinjector, for migraine patients

Elimor Brand-Schieber,1 Sagar Munjal,1 Rajesh Kumar,1 Anthony D Andre,2 Will Valladao,2 Margarita Ramirez2 1Dr. Reddy’s Laboratories Inc., Princeton, NJ, 2Interface Analysis Associates, Saratoga, CA, USA Background: Migraine pain relief is reported by more than 50% of patients who receive low dose (3 mg) of sumatriptan. Currently, there is no two-step autoinjector of low-dose sumatriptan available on the market for acute migraine treatment. To fulfill this need, a fully assembled, single-dose, subcutaneous autoinjector (sumatriptan 3 mg; product-code DFN-11) was developed. The device allows for injection with a simple two-step, push-to-inject process and provides feedback of the injection activation, progress, and completion.Objective: To determine if DFN-11 autoinjector can be used correctly and safely by migraine patients.Methods and participants: A human factors validation study was conducted with 45 migraine patients (30 oral-only medications users; 15 injectable sumatriptan users) who performed one unaided simulated injection. Two days prior, half the oral participants were briefly trained. All others were only given the device to inspect and written instructions to review. No injections were performed during the initial session. All participants received written instructions at the injection session.Results: All participants (45/45; 100%) performed the injection without any errors. Objective measures included device removal from packaging, cap removal, expiration date check, ­inspection of fluid in window, identification of allowable injection site, proper device positioning, dose confirmation, and device disposal. All participants (45/45; 100%) reported no difficulty administering the injection and no concerns about using the autoinjector during a severe migraine onset.Conclusion: The results showed that the DFN-11 autoinjector can be used with safe handling without patterns of confusion, failures, high-risk errors, wet injections, or patient safety risks. The DFN-11 autoinjector was validated to be used correctly and safely by migraine patients, whether they were injection experienced, unexperienced, trained, or self-trained. Keywords: triptan, pain relief, subcutaneous injection, preference, usabilit