Regulation of activity of P2X7 receptor by its splice variants in cultured mouse astrocytes

Of purinergic receptors, P2X7 receptor (P2X7R, defined as a full‐length receptor) has unique characteristics, and its activation leads to ion channel activity and pore formation, causing cell death. Previously, we demonstrated that P2X7R expressed by nonstimulated astrocyte cultures obtained from SJL‐strain mice exhibits constitutive activation, implying its role in maintenance of cellular homeostasis. To obtain novel insights into its physiological roles, we examined whether constitutive activation of P2X7R is regulated by expression of its splice variants in such resting astrocytes, and whether their distinct expression profiles in different mouse strains affect activation levels of astrocytic P2X7Rs. In SJL‐ and ddY‐mouse astrocytes, spontaneous YO‐PRO‐1 uptake, an indicator of pore activity of P2X7R, was detected, but the uptake by the formers was significantly greater than that by the latter. Between the two mouse strains, there was a difference in their sensitivity of YO‐PRO‐1 uptake to antagonists, but not in the expression levels and sequences of P2X7R and pannexin‐1. Regarding expression of splice variants of P2X7R, expression of P2X7R variant‐3 (P2X7R‐v3) and ‐4 (P2X7R‐v4), but not variant‐2 and ‐k, was lower in SJL‐mouse astrocytes than in ddY‐mouse ones. On transfection of P2X7R‐v3 and ‐v4 into SJL‐mouse astrocytes, the pore activity was attenuated as in the case of the HEK293T cell‐expression system. These findings demonstrate that basal activity of P2X7R expressed by resting astrocytes is negatively regulated by P2X7R‐v3 and ‐v4, and that their distinct expression profiles result in the different activation levels of astrocytic P2X7Rs in different mouse strains. GLIA 2014;62:440–451     

The role of hypothalamic estrogen receptors in metabolic regulation

Estrogens regulate key features of metabolism, including food intake, body weight, energy expenditure, insulin sensitivity, leptin sensitivity, and body fat distribution. There are two ‘classical’ estrogen receptors (ERs): estrogen receptor alpha (ERS1) and estrogen receptor beta (ERS2). Human and murine data indicate ERS1 contributes to metabolic regulation more so than ESR2. For example, there are human inactivating mutations of ERS1 which recapitulate aspects of the metabolic syndrome in both men and women. Much of our understanding of the metabolic roles of ERS1 was initially uncovered in estrogen receptor α-null mice (ERS1^−/−); these mice display aspects of the metabolic syndrome, including increased body weight, increased visceral fat deposition and dysregulated glucose intolerance. Recent data further implicate ERS1 in specific tissues and neuronal populations as being critical for regulating food intake, energy expenditure, body fat distribution and adipose tissue function. This review will focus predominantly on the role of hypothalamic ERs and their critical role in regulating all aspects of energy homeostasis and metabolism.     

Electrical impedance myography for the in vivo and ex vivo assessment of muscular dystrophy (mdx) mouse muscle

Introduction: Sensitive, non‐invasive techniques are needed that can provide biomarkers of disease status and the effects of therapy in muscular dystrophy. Methods: We evaluated electrical impedance myography (EIM) to serve in this role by studying 2‐month‐old and 18‐month‐old mdx and wild‐type (WT) animals (10 animals in each of 4 groups). Results: Marked differences were observed in EIM values between mdx and WT animals; the differences were more pronounced between the older age groups (e.g., reactance of 92.6 ±4.3 Ω for mdx animals vs. 130 ± 4.1 Ω for WT animals, P < 0.001). In addition, in vivo EIM parameters correlated significantly with the extent of connective tissue deposition in the mdx animals. Conclusions: EIM has the potential to serve as a valuable non‐invasive method for evaluating muscular dystrophy. It can be a useful biomarker to assist with therapeutic testing in both pre‐clinical and clinical studies. Muscle Nerve 49 : 829–835, 2014     

Characterization of the mitofusin 2 R94W mutation in a knock‐in mouse model

Charcot‐Marie‐Tooth disease (CMT) comprises a group of heterogeneous peripheral axonopathies affecting 1 in 2,500 individuals. As mutations in several genes cause axonal degeneration in CMT type 2, mutations in mitofusin 2 (MFN2) account for approximately 90% of the most severe cases, making it the most common cause of inherited peripheral axonal degeneration. MFN2 is an integral mitochondrial outer membrane protein that plays a major role in mitochondrial fusion and motility; yet the mechanism by which dominant mutations in this protein lead to neurodegeneration is still not fully understood. Furthermore, future pre‐clinical drug trials will be in need of validated rodent models. We have generated a Mfn2 knock‐in mouse model expressing Mfn2^R94W, which was originally identified in CMT patients. We have performed behavioral, morphological, and biochemical studies to investigate the consequences of this mutation. Homozygous inheritance leads to premature death at P1, as well as mitochondrial dysfunction, including increased mitochondrial fragmentation in mouse embryonic fibroblasts and decreased ATP levels in newborn brains. Mfn2^R94W heterozygous mice show histopathology and age‐dependent open‐field test abnormalities, which support a mild peripheral neuropathy. Although behavior does not mimic the severity of the human disease phenotype, this mouse can provide useful tissues for studying molecular pathways associated with MFN2 point mutations.     

Experimental and finite element analysis of dynamic loading of the mouse forearm

Bone formation is reported to initiate in osteocytes by mechanotransduction due to dynamic loading of bone. The first step towards this is to characterize the dynamic strain fields in the overall bone. Here, the previously developed mouse forearm ulna‐radius model, subjected to static loading, has been further enhanced by incorporating a loading cap and applying a cyclic dynamic load to more closely approximate experimental biological conditions. This study also incorporates data obtained from strain gauging both the ulna and radius simultaneously. Based on separate experiments, the elastic modulus of the ulna and radius were determined to be 13.8 and 9.9 GPa, respectively. Another novel aspect of the numerical model is the inclusion of the interosseous membrane in the FE model with membrane stiffness ranging from 5–15 N/mm that have been found to give strain values closer to that from the experiments. Interestingly, the inclusion of the interosseous membrane helped to equalize the peak strain magnitudes in the ulna and radius (～1800 at 2 N load and ～3200 at 3.5 N), which was also observed experimentally. This model represents a significant advance towards being able to simulate through FE analysis the strain fields generated in vivo upon mechanical loading of the mouse forearm. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:1580–1588, 2014.