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Hibernation is a natural model of extreme physiology in a mammal. Throughout winter, small hibernators repeatedly undergo rapid, dramatic swings in body temperature, perfusion, and oxygen delivery. To gain insight into the molecular mechanisms that support homeostasis despite the numerous challenges posed by this dynamic physiology, we collected 13-lined ground squirrel adrenal glands from at least five individuals representing six key timepoints across the year using body temperature telemetry. Differentially expressed genes were identified using RNA-seq, revealing both strong seasonal and torpor-arousal cycle effects on gene expression. Two novel findings emerge from this study. First, transcripts encoding multiple genes involved in steroidogenesis decreased seasonally. Taken together with morphometric analyses, the data are consistent with preservation of mineralocorticoids but suppression of glucocorticoid and androgen output throughout winter hibernation. Second, a temporally orchestrated, serial gene expression program unfolds across the brief arousal periods. This program initiates during early rewarming with the transient activation of a set of immediate early response (IER) genes, comprised of both transcription factors and the RNA degradation proteins that assure their rapid turnover. This pulse in turn activates a cellular stress response program to restore proteostasis comprised of protein turnover, synthesis, and folding machinery. These and other data support a general model for gene expression across the torpor-arousal cycle that is facilitated in synchrony with whole body temperature shifts; induction of the immediate early response upon rewarming activates a proteostasis program followed by a restored tissue-specific gene expression profile enabling renewal, repair, and survival of the torpid state. [ABSTRACT FROM AUTHOR]

The extensive oxygen gradient between the air we breathe (PO2 ~21 kPa) and its ultimate distribution within mitochondria (as low as ~0.5-1 kPa) is testament to the efforts expended in limiting its inherent toxicity. It has long been recognized that cell culture undertaken under room air conditions falls short of replicating this protection in vitro. Despite this, difficulty in accurately determining the appropriate O2 levels in which to culture cells, coupled with a lack of the technology to replicate and maintain a physiological O2 environment in vitro, has hindered addressing this issue thus far. In this review, we aim to address the current understanding of tissue PO2 distribution in vivo and summarize the attempts made to replicate these conditions in vitro. The state-of-the-art techniques employed to accurately determine O2 levels, as well as the issues associated with reproducing physiological O2 levels in vitro, are also critically reviewed. We aim to provide the framework for researchers to undertake cell culture under O2 levels relevant to specific tissues and organs. We envisage that this review will facilitate a paradigm shift, enabling translation of findings under physiological conditions in vitro to disease pathology and the design of novel therapeutics. [ABSTRACT FROM AUTHOR]

Much innovation is currently aimed at improving the number, density, and geometry of electrodes on extracellular multielectrode arrays for in vivo recording of neural activity in the mammalian brain. To choose a multielectrode array configuration for a given neuroscience purpose, or to reveal design principles of future multielectrode arrays, it would be useful to have a systematic way of evaluating the spike recording capability of such arrays. We describe an automated system that performs robotic patch-clamp recording of a neuron being simultaneously recorded via an extracellular multielectrode array. By recording a patch-clamp data set from a neuron while acquiring extracellular recordings from the same neuron, we can evaluate how well the extracellular multielectrode array captures the spiking information from that neuron. To demonstrate the utility of our system, we show that it can provide data from the mammalian cortex to evaluate how the spike sorting performance of a close-packed extracellular multielectrode array is affected by bursting, which alters the shape and amplitude of spikes in a train. We also introduce an algorithmic framework to help evaluate how the number of electrodes in a multielectrode array affects spike sorting, examining how adding more electrodes yields data that can be spike sorted more easily. Our automated methodology may thus help with the evaluation of new electrode designs and configurations, providing empirical guidance on the kinds of electrodes that will be optimal for different brain regions, cell types, and species, for improving the accuracy of spike sorting. [ABSTRACT FROM AUTHOR]

Mammalian hibernators, such as golden-mantled ground squirrels (Callospermophilus lateralis; GMGS), cease to feed while reducing metabolic rate and body temperature during winter months, surviving exclusively on endogenous fuels stored before hibernation. We hypothesized that mitochondria, the cellular sites of oxidative metabolism, undergo tissue-specific seasonal adjustments in carbohydrate and fatty acid utilization to facilitate or complement this remarkable phenotype. To address this, we performed high-resolution respirometry of mitochondria isolated from GMGS liver, heart, skeletal muscle, and brown adipose tissue (BAT) sampled during summer (active), fall (prehibernation), and winter (hibernation) seasons using multisubstrate titration protocols. Mitochondrial phospholipid composition was examined as a postulated intrinsic modulator of respiratory function across tissues and seasons. Respirometry revealed seasonal variations in mitochondrial oxidative phosphorylation capacity, substrate utilization, and coupling efficiency that reflected the distinct functions and metabolic demands of the tissues they support. A consistent finding across tissues was a greater influence of fatty acids (palmitoylcarnitine) on respiratory parameters during the prehibernation and hibernation seasons. In particular, fatty acids had a greater suppressive effect on pyruvate-supported oxidative phosphorylation in heart, muscle, and liver mitochondria and enhanced uncoupled respiration in BAT and muscle mitochondria in the colder seasons. Seasonal variations in the mitochondrial membrane composition re- flected changes in the supply and utilization of polyunsaturated fatty acids but were generally mild and inconsistent with functional variations. In conclusion, mitochondria respond to seasonal variations in physical activity, temperature, and nutrient availability in a tissuespecific manner that complements circannual shifts in the bioenergetic and thermoregulatory demands of mammalian hibernators. [ABSTRACT FROM AUTHOR]

During hibernation, thirteen-lined ground squirrels (Ictidomys tridecemlineatus) regularly cycle between bouts of torpor and interbout arousal (IBA). Most of the brain is electrically quiescent during torpor but regains activity quickly upon arousal to IBA, resulting in extreme oscillations in energy demand during hibernation. We predicted increased functional capacity of brain mitochondria during hibernation compared with spring to accommodate the variable energy demands of hibernation. To address this hypothesis, we examined mitochondrial bioenergetics in the ground squirrel brain across three time points: spring (SP), torpor (TOR), and IBA. Respiration rates of isolated brain mitochondria through complex I of the electron transport chain were more than twofold higher in TOR and IBA than in SP (P < 0.05). We also found a 10% increase in membrane potential between hibernation and spring (P < 0.05), and that proton leak was lower in TOR and IBA than in SP. Finally, there was a 30% increase in calcium loading in SP brain mitochondria compared with TOR and IBA (P < 0.01). To analyze brain mitochondrial abundance between spring and hibernation, we measured the ratio of copy number in a mitochondrial gene (ND1) vs. a nuclear gene (B2M) in frozen cerebral cortex samples. No significant differences were observed in DNA copies between SP and IBA. These data show that brain mitochondrial bioenergetics are not static across the year and suggest that brain mitochondria function more effectively during the hibernation season, allowing for rapid production of energy to meet demand when extreme physiological changes are occurring. [ABSTRACT FROM AUTHOR]

Mammalian hibernators adapt to prolonged periods of immobility, hypometabolism, hypothermia, and oxidative stress, each capable of reducing bone marrow activity. In this study bone marrow transcriptomes were compared among thirteen-lined ground squirrels collected in July, winter torpor, and winter interbout arousal (IBA). The results were consistent with a suppression of acquired immune responses, and a shift to innate immune responses during hibernation through higher complement expression. Consistent with the increase in adipocytes found in bone marrow of hibernators, expression of genes associated with white adipose tissue are higher during hibernation. Genes that should strengthen the bone by increasing extracellular matrix were higher during hibernation, especially the collagen genes. Finally, expression of heat shock proteins were lower, and cold-response genes were higher, during hibernation. No differential expression of hematopoietic genes involved in erythrocyte or megakaryocyte production was observed. This global view of the changes in the bone marrow transcriptome over both short term (torpor vs. IBA) and long term (torpor vs. July) hypothermia can explain several observations made about circulating blood cells and the structure and strength of the bone during hibernation. [ABSTRACT FROM AUTHOR]

Mammalian spermatogenesis requires a stem cell pool, a period of amplification of cell numbers, the completion of reduction division to haploid cells (meiosis), and the morphological transformation of the haploid cells into spermatozoa (spermiogenesis). The net result of these processes is the production of massive numbers of spermatozoa over the reproductive lifetime of the animal. One study that utilized homogenization-resistant spermatids as the standard determined that human daily sperm production (dsp) was at 45 million per day per testis (60). For each human that means ~1,000 sperm are produced per second. A key to this level of gamete production is the organization and architecture of the mammalian testes that results in continuous sperm production. The seemingly complex repetitious relationship of cells termed the "cycle of the seminiferous epithelium" is driven by the continuous commitment of undifferentiated spermatogonia to meiosis and the period of time required to form spermatozoa. This commitment termed the A to A1 transition requires the action of retinoic acid (RA) on the undifferentiated spermatogonia or prospermatogonia. In stages VII to IX of the cycle of the seminiferous epithelium, Sertoli cells and germ cells are influenced by pulses of RA. These pulses of RA move along the seminiferous tubules coincident with the spermatogenic wave, presumably undergoing constant synthesis and degradation. The RA pulse then serves as a trigger to commit undifferentiated progenitor cells to the rigidly timed pathway into meiosis and spermatid differentiation. [ABSTRACT FROM AUTHOR]

Manganese superoxide dismutase (MnSOD) is considered a critical component of the antioxidant systems that protect against oxidative damage. We are interested in the role of oxidative stress in bladder detrusor smooth muscle (SM) in different disease states. In this study, we generated an inducible, SM-specific Sod2-/- mouse model to investigate the effects of MnSOD depletion on the function of the bladder. We crossbred floxed Sod2 (Sod2lox/lox) mice with mice containing heterozygous knock-in of a gene encoding a tamoxifen-activated Cre recombinase in the SM22α promoter locus [SM-CreERT2(ki)Cre/+]. We obtained Sod2lox/lox,SM-CreERT2(ki)Cre/+ mice and injected 8-wk-old males with 4-hydroxytamoxifen to induce Cre-mediated excision of the floxed Sod2 allele. Twelve weeks later, SM-specific deletion of Sod2 and depletion of MnSOD were confirmed by polymerase chain reaction, immunoblotting, and immunohistochemistry. SM-specific Sod2-/- mice exhibited normal growth with no gross abnormalities. A significant increase in nitrotyrosine concentration was found in bladder SM tissue of SM-specific Sod2-/- mice compared with both wild-type mice and Sod2+/+, SM-CreERT2(ki)Cre/+ mice treated with 4-hydroxytamoxifen. Assessment of 24-h micturition in SM-specific Sod2-/- mice revealed significantly higher voiding frequency compared with both wild-type and SM-specific Cre controls. Conscious cystometry revealed significantly shorter intercontraction intervals and lower functional bladder capacity in SM-specific Sod2-/- mice compared with wild-type mice. This novel model can be used for exploring the mechanistic role of oxidative stress in organs rich in SM in different pathological conditions. [ABSTRACT FROM AUTHOR]

Little is known about steroidal control of thirst- and salt-appetite behaviors of mice. The current study investigates effects of fludrocortisone acetate (FCA), a steroid with potent glucocorticoid and mineralocorticoid effects, on thirst- and salt-appetite responses of C57BL/6 mice. Treatment with FCA produced dose-dependent (5, 10, and 25 mg/kg) increases in both magnitude and duration of water and sodium intake. Chronic elevation of water and saline intake was achieved with daily injections of FCA. Daily injection of FCA, when only 0.9% saline was available, produced a remarkably rapid increase in saline intake. A single injection of FCA stimulated brisk diuresis and natriuresis in fluid-restricted animals. This work is the first to demonstrate copious water drinking by mice in response to FCA. The results are discussed in terms of the possibility that the renal effects of FCA promote increases in water and sodium turnover and thereby, increases in water and sodium ingestion. [ABSTRACT FROM AUTHOR]

Mammals are often considered to be masters of homeostasis, with the ability to maintain a constant internal milieu, despite marked changes in the environment; however, many species exhibit striking physiological and biochemical plasticity in the face of environmental fluctuations. Here, we review metabolic depression and body temperature fluctuation in mammals, with a focus on the extreme example of hibernation in small-bodied eutherian species. Careful exploitation of the phenotypic plasticity of mammals with metabolic flexibility may provide the key to unlocking the molecular secrets of orchestrating and surviving reversible metabolic depression in less plastic species, including humans. [ABSTRACT FROM AUTHOR]

Overnutrition/obesity predisposes individuals, particularly women, to diastolic dysfunction (DD), an independent predictor of future cardiovascular disease. We examined whether low-dose spironolactone (Sp) prevents DD associated with consumption of a Western Diet (WD) high in fat, fructose, and sucrose. Female C57BL6J mice were fed a WD with or without Sp (1 mg·kg-1·day-1). After 4 mo on the WD, mice exhibited increased body weight and visceral fat, but similar blood pressures, compared with control diet-fed mice. Sp prevented the development of WD-induced DD, as indicated by decreased isovolumic relaxation time and an improvement in myocardial performance (

Neurons throughout the mammalian central auditory pathway respond selectively to stimulus frequency and amplitude, and some are also selective for stimulus duration. First found in the auditory midbrain or inferior colliculus (IC), these duration-tuned neurons (DTNs) provide a potential neural mechanism for encoding temporal features of sound. In this study, we investigated how having an additional neural response filter, one selective to the duration of an auditory stimulus, influences frequency tuning and neural organization by recording single-unit responses and measuring the dorsal-ventral position and spectral-temporal tuning properties of auditory DTNs from the IC of the awake big brown bat (Eptesicus fuscus). Like other IC neurons, DTNs were tonotopically organized and had either V-shaped, U-shaped, or O-shaped frequency tuning curves (excitatory frequency response areas). We hypothesized there would be an interaction between frequency and duration tuning in DTNs, as electrical engineering theory for resonant filters dictates a trade-off in spectral-temporal resolution: sharp tuning in the frequency domain results in poorer resolution in the time domain and vice versa. While the IC is a more complex signal analyzer than an electrical filter, a similar operational trade-off could exist in the responses of DTNs. Our data revealed two patterns of spectrotemporal sensitivity and spatial organization within the IC: DTNs with sharp frequency tuning and broad duration tuning were located in the dorsal IC, whereas cells with wide spectral tuning and narrow temporal tuning were found in the ventral IC. [ABSTRACT FROM AUTHOR]

The p66Shc adapter protein regulates the morphogenesis and epithelial maturation of fetal mouse lungs. Am J Physiol Lung Cell Mol Physiol 306: L316-L325, 2014. First published December 27, 2013; doi:10.1152/ajplung.00062.2013.--Many signaling pathways are mediated by Shc adapter proteins that, in turn, are expressed as three isoforms with distinct functions. The p66Shc isoform antagonizes proliferation, regulates oxidative stress, and mediates apoptosis. It is highly expressed in the canalicular but not the later stages of mouse lung development, and its expression persists in bronchopulmonary dysplasia, a chronic disease associated with premature birth. These observations suggest that p66Shc has a developmental function. However, constitutive p66Shc deletion yields no morphological phenotype, and the structure of the Shc gene precludes its inducible deletion. To elucidate its function in lung development, we transfected p66Shc or nonsilencing small-interfering RNA (siRNA) into the epithelia of embryonic day 11 mouse lungs that were then cultured for 3 days and analyzed morphometrically. To assess cellular proliferation and epithelial differentiation, lung explants were immunostained and immunoblotted for p66Shc, proliferating cell nuclear antigen (PCNA), the proximal airway differentiation antigens Clara cell 10-kDa protein (CC10) and thyroid transcription factor (TTF)-1, and the alveolar surfactant proteins (SP)-A, -B, and -C. Explants transfected with nonsilencing siRNA demonstrated specific epithelial uptake and normal morphological development relative to uninjected controls. In contrast, transfection with p66Shc siRNA significantly increased lumenal cross-sectional areas, decreased branching, and increased epithelial proliferation (P < 0.05 for all). Relative to controls, the expression of SP-B, SP-C, CC10, and TTF-1 was decreased by p66Shc knockdown. SP-A was not expressed in either control or treated lungs. These data suggest that p66Shc attenuates epithelial proliferation while promoting both distal and proximal epithelial maturation. [ABSTRACT FROM AUTHOR]

Superoxide dismutase 2 (SOD-2) is synthesized in the cytosol and imported into the mitochondrial matrix, where it is activated and functions as the primary antioxidant for cellular respiration. The specific mechanisms that target SOD-2 to the mitochondria remain unclear. We hypothesize that inducible heat shock protein 70 (iHSP70) targets SOD-2 to the mitochondria via a mechanism facilitated by ATP, and this process is impaired in persistent pulmonary hypertension of the newborn (PPHN). We observed that iHSP70 interacts with SOD-2 and targets SOD-2 to the mitochondria. Interruption of iHSP70-SOD-2 interaction with 2-phenylethylenesulfonamide- μ (PFT-μ, a specific inhibitor of substrate binding to iHSP70 COOH terminus) and siRNA-mediated knockdown of iHSP70 expression disrupted SOD-2 transport to mitochondria. Increasing intracellular ATP levels by stimulation of respiration with CaCl2 facilitated the mitochondrial import of SOD-2, increased SOD-2 activity, and decreased the mitochondrial superoxide (O2 ·-) levels in PPHN pulmonary artery endothelial cells (PAEC) by promoting iHSP70-SOD-2 dissociation at the outer mitochondrial membrane. In contrast, oligomycin, an inhibitor of mitochondrial ATPase, decreased SOD-2 expression and activity and increased O2 ·- levels in the mitochondria of control PAEC. The basal ATP levels and degree of iHSP70-SOD-2 dissociation were lower in PPHN PAEC and lead to increased SOD-2 degradation in cytosol. In normal pulmonary arteries (PA), PFT-μ impaired the relaxation response of PA rings in response to nitric oxide (NO) donor, S-nitroso-N-acetyl-penicillamine. Pretreatment with Mito-Q, a mitochondrial targeted O2 ·- scavenger, restored the relaxation response in PA rings pretreated with PFT-μ. Our observations suggest that iHSP70 chaperones SOD-2 to the mitochondria. Impaired SOD-2-iHSP70 dissociation decreases SOD-2 import and contributes to mitochondrial oxidative stress in PPHN. [ABSTRACT FROM AUTHOR]

Excess iron causes oxidative stress through hydroxyl-radical production via Fenton/ Haber-Weiss reactions. Recently, body iron reduction has been found to ameliorate diabetes. In the present study, we examined the protective effect of dietary iron restriction against diabetic nephropathy in the db/db mouse model of diabetic nephropathy using db/m mice as controls. The db/db mice were divided into two groups and fed a normal diet (ND) or a low-iron diet (LID). Increasing urinary albumin excretion was observed in the ND db/db mice, but this was suppressed in db/db mice with LID. Histologically, the db/db mice in the ND group had increased glomerular volume and mesangial area compared with the LID group. Augmented deposition of extracellular matrixes was decreased in db/db mice with LID. In terms of oxidative stress, increased superoxide production observed in the kidneys of the ND db/db mice was diminished in the LID group. NADPH oxidase activity and renal expression of NADPH oxidase components p22phox and NADPH oxidase 4 (NOX4) were augmented in the ND group, and this was abolished by LID. There were no differences in expression of renal iron importers, transferrin receptor, or divalent metal transporter-1 between db/m mice and db/db mice. The level of ferroportin, an iron exporter, increased in the kidneys of the db/db mice. Urinary iron excretion was significantly higher in ND db/db mice and was reduced in the LID group. These findings suggest that dietary iron restriction exerts a preventive effect on the progression of diabetic nephropathy partly due to the reduction of oxidative stress. [ABSTRACT FROM AUTHOR]

Deoxycorticosterone salt (DOC-salt) hypertension-induced renal damage is enhanced in α-calcitonin gene-related peptide (α-CGRP) knockout (KO) compared with wild-type (WT) mice. However, since the α-CGRP KO mice have a 15-20 mmHg higher baseline mean arterial pressure (MAP) than WT mice, they also have a higher MAP than WT mice throughout the course of DOC-salt hypertension. To determine the mechanism by which the absence of α-CGRP enhances hypertension-induced renal damage, DOC-salt hypertension was induced in telemetry probe implanted α-CGRP KO and WT mice. To equalize the blood pressure (BP) to that of DOC-salt WT mice, an additional group of DOC-salt α-CGRP KO mice was given 0.025% hydralazine to drink. The DOC-salt protocol increased the final MAP in α-CGRP KO mice to 155 ± 6 mmHg and in WT mice to 140 ± 5 mmHg. The MAP of the hydralazine-treated DOC-salt α-CGRP KO mice was 139 ± 6 mmHg. Urinary excretion of microalbumin and isoprostane, a marker for oxidative stress, was increased, and creatinine clearance was decreased in DOC-salt α-CGRP KO compared with DOC-salt WT mice. Equalization of the MAP in DOC-salt α-CGRP KO to that of DOC-salt WT mice did not significantly improve these parameters. Renal macrophage infiltration; desmin, a marker of podocyte damage; and the inflammatory cytokines TNF-α and IFN-ɣ and the chemokines monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1α (MIP-1α) were increased in DOC-salt α-CGRP KO mice and were not reduced by hydralazine treatment. However, BP equalization did improve the renal histopathological damage, as determined by light microscopy. Therefore, in DOC-salt hypertension in mice, the mechanism(s) of the renal protective effects of α-CGRP are both BP independent and BP dependent. [ABSTRACT FROM AUTHOR]

Immune cell infiltration of the kidney is a constant feature in salt-sensitive hypertension (SSHTN). We evaluated the relationship between the renal inflammation and pressure natriuresis in the model of SSHTN that results from transient oral administration of Nω-nitro-L-arginine methyl ester (L-NAME). Pressure natriuresis was determined in Wistar rats that received 4 wk of a high-salt (4% NaCl) diet, starting 1 wk after stopping L-NAME, which was administered alone (SSHTN group, n = 17) or in association with mycophenolate mofetil (MMF; MMF group, n 15). The administration of MMF in association with L-NAME is known to prevent the subsequent development of SSHTN. Control groups received a high (n = 12)- and normal (0.4%)-salt diet (n = 20). Rats with SSHTN had increased expression of inflammatory cytokines and oxidative stress. The severity of hypertension correlated directly (P ω 0.0001) with the number of tubulointerstitial immune cells and angiotensin II-expressing cells. Pressure natriuresis was studied at renal arterial pressures (RAPs) of 90, 110, 130, and 150 mmHg. Glomerular filtration rate was similar and stable in all groups, and renal blood flow was decreased in the SSHTN group. Significantly decreased natriuresis (P 0.05) was found in the SSHTN group at RAPs of 130 and 150 mmHg, and there was an inverse correlation (P ω 0.01) between the urinary sodium excretion and the number of tubulointerstitial inflammatory cells (lymphocytes and macrophages) and cells expressing angiotensin II. We conclude that tubulointerstitial inflammation plays a key role in the impairment of pressure natriuresis that results in salt-dependent hypertension in this experimental model. [ABSTRACT FROM AUTHOR]

A growing portion of premedical curricula is being devoted to the study of physiological mechanisms underlying animal behavior. In the present article, I describe an activity centered around a classic Journal of Neurophysiology paper by Edward V. Evarts that lays the foundation for students to investigate common behavioral and physiological techniques used to study motor control in primates. Students will leave this activity being able to 1) critically assess behavioral, electromyographic, and single unit (extracellular) neurophysiological data typically acquired by behavioral neurophysiologists; 2) provide physiological evidence that the primate precentral gyrus (primary motor cortex) controls voluntary movements of the wrist; 3) intelligently discuss hypotheses concerning the role of the primary motor cortex in the generation of movement in mammalian species; and 4) discuss the ethical implications of using mammalian species as model organisms. The skills and background knowledge gained in this activity lay the platform for advanced study of scientific investigations into sensory, motor, and cognitive processes in undergraduate, graduate, or medical school curricula. [ABSTRACT FROM AUTHOR]

Transverse (t) tubules are surface membrane invaginations that are present in all mammalian cardiac ventricular cells. The apposition of L-type Ca2+ channels on t tubules with the sarcoplasmic reticulum (SR) constitutes a "calcium release unit" and allows close coupling of excitation to the rise in systolic Ca2+. T tubules are virtually absent in the atria of small mammals, and therefore Ca2+ release from the SR occurs initially at the periphery of the cell and then propagates into the interior. Recent work has, however, shown the occurrence of t tubules in atrial myocytes from sheep. As in the ventricle, Ca2+ release in these cells occurs simultaneously in central and peripheral regions. T tubules in both the atria and the ventricle are lost in disease, contributing to cellular dysfunction. The aim of this study was to determine if the occurrence of t tubules in the atrium is restricted to sheep or is a more general property of larger mammals including humans. In atrial tissue sections from human, horse, cow, and sheep, membranes were labeled using wheat germ agglutinin. As previously shown in sheep, extensive t-tubule networks were present in horse, cow, and human atrial myocytes. Analysis shows half the volume of the cell lies within 0.64 ± 0.03, 0.77 ± 0.03, 0.84 ± 0.03, and 1.56 ± 0.19 µm of t-tubule membrane in horse, cow, sheep, and human atrial myocytes, respectively. The presence of t tubules in the human atria may play an important role in determining the spatio-temporal properties of the systolic Ca2+ transient and how this is perturbed in disease. [ABSTRACT FROM AUTHOR]

The article focuses on the review of mammalian muscle fiber heterogeneity and the integrative function and structure of various types of skeletal muscle fiber. It says the heterogeneity of muscle fibers is the basis for flexibility which allows muscle to be used for various tasks. It discusses the major type of fibers composed of distinct myosin and the minor fiber types with restricted distribution. Moreover, selected signaling pathways and adaptive changes in muscle plasticity is also cited.

Resetting the circadian clock leads to well being and increased life span, whereas clock disruption is associated with aging and morbidity. Increased longevity and improved health can be achieved by different feeding regimens that reset circadian rhythms and may lead to better synchrony in metabolism and physiology. This review focuses on recent findings concerning the relationships between circadian rhythms, aging attenuation, and life-span extension in mammals. [ABSTRACT FROM AUTHOR]

This study provides evidence that peer marking can be a reliable tool for assessing laboratory reports in large cohorts. It was conducted over a 4-yr period with first-year undergraduates (∼180 students/cohort) taking a mammalian physiology course, but the procedure adopted would be applicable to any other laboratory-based discipline. The process was found to be efficient in staff time, enabling a summative practical report to be marked in <1 h (<5% of the time that had previously been required for staff marking), facilitating rapid feedback to students on their performance. When samples of the peer-assessed reports were marked by a single member of staff, there was excellent correlation between peer and staff marks (r = 0.96-0.98), although peer-awarded marks exceeded staff marks by an average of 2.5-3.0%. The validity of peer marking was independent of both the sex of the marker and the staff score awarded to the marker for the same piece of work. Feedback from students was largely positive; they reported that the procedure adopted was effective in increasing their understanding of the underlying physiology and contributed to their understanding of best practice in presenting a laboratory report. Seventy percent of students agreed that it was acceptable for peer assessment to contribute a small (up to 5%) component of the overall mark for the course. The results are discussed in relation to other reports of peer marking, particularly when used to assess an academic product or process in a scientific discipline. [ABSTRACT FROM AUTHOR]

The optical recording of light-evoked activity in populations of neurons in the mammalian retina offers several benefits over the use of multielectrode arrays. However, population imaging has been hindered by the effective loading of synthetic fluorescent indicators, especially in the mature tissue. We have therefore developed an electroporation method to label the complete ganglion cell layer of the adult mammalian retina. We optimized the protocol such that the retina recovers from electroporation and generates responses to visual stimuli. The method can be used with a diverse set of indicators with a range of affinities and emission wavelengths. It therefore can be combined with transgenic animals expressing fluorescent markers to target specific neuronal types. Importantly, the ganglion cell layer remains accessible for subsequent intracellular recording and morphological identification. [ABSTRACT FROM AUTHOR]

The article examines the neurophysiological function of cerebral cortex in awake mammals. It discusses the cognitive process in the brain region, involving the synchronization and stochastic neural activity of gamma and theta oscillations using electroencephalogram. It reveals the relation between the irregularity of neural activity and mechanism of the cortical rhythms. It also covers the association of synchronization dysfunction in mental disorders such as schizophrenia and autism.

The article focuses on the entrainment mechanisms and the physiology of the circadian rhythms of mammals. It explores the retinohypothalamic communication of circadian rhythms, features of circadian system, and the chronobiology of the circadian rhythms. It discusses and analyzes the endogenous biological oscillators like the master clock situated in the hypothalamic supraschiasmatic nuclei (SCN) which controls the circadian rhythms, and the clock gene expression.

The objective of this study was to determine whether the previously observed effects of photoperiod on body weight in Siberian hamsters were due to changes in the daily patterns of locomotor activity, energy expenditure, and/or feeding behavior. Adult males were monitored through a seasonal cycle using an automated comprehensive laboratory animal monitoring system (CLAMS). Exposure to a short-day photoperiod (SD; 8:16-h light-dark cycle) induced a significant decline in body weight, and oxygen consumption (Vo2), carbon dioxide production (Vco2), and heat production all decreased reaching a nadir by 16 wk of SD. Clear daily rhythms in locomotor activity, Vo2, and Vco2 were observed at the start of the study, but these all progressively diminished after prolonged exposure to SD. Rhythms in feeding behavior were also detected initially, reflecting an increase in meal frequency but not duration during the dark phase. This rhythm was lost by 8 wk of SD exposure such that food intake was relatively constant across dark and light phases. After 18 wk in SD, hamsters were transferred to a long-day photoperiod (LD; 16:8-h light-dark cycle), which induced significant weight gain. This was associated with an increase in energy intake within 2 wk, while Vo2, Vco2, and heat production all increased back to basal levels. Rhythmicity was reestablished within 4 wk of reexposure to long days. These results demonstrate that photoperiod impacts on body weight via complex changes in locomotor activity, energy expenditure, and feeding behavior, with a striking loss of daily rhythms during SD exposure [ABSTRACT FROM AUTHOR]

Hypoxia is an important ecological, evolutionary, and biomedical stressor. While physiological acclimatization of mammals to hypoxia is well studied, the variation in gene expression that underlies acclimatization is not well studied. We acclimatized inbred mice for 32 days to hypoxic conditions that simulated altitudes of 1400, 3000, and 4500 m. We used oligonucleotide microarrays to measure changes in steady-state abundance of mRNA in the livers of these mice. Mice exposed to more severe hypoxia (simulated altitude of 4500 m) were smaller in mass and had higher hematocrit than mice exposed to less severe hypoxia. ANOVA and false discovery rate tests indicated that 580 genes were significantly differentially expressed in response to chronic hypoxia. Few of these 580 genes have previously been reported to respond to hypoxia. In contrast, many of these 580 genes belonged to same functional groups typically respond to acute hypoxia. That is, both chronic and acute hypoxia elicit changes in transcript abundance for genes involved in angiogenesis, glycolysis, lipid metabolism, carbohydrate metabolism, and protein amino acid phosphorylation, but the particular genes affected by the two types of hypoxia were mostly different. Numerous genes affecting the immune system were differentially expressed in response to chronic hypoxia, which supports recently proposed hypotheses that link immune function and hypoxia. Furthermore, our results discovered novel elevated mRNA abundance of genes involved in hematopoiesis and oxygen transport not reported previously, but consistent with extreme hematocrits found in hypoxic mice. [ABSTRACT FROM AUTHOR]

The article focuses on ventilatory response exhibited by mammals to counter anoxemia. It explores the elements that control breathing of fetal and neonatal mammals governed by the contraction of respiratory muscles following a rhythm. It also notes that carotid bodies such as glomus cells and sustentacular cells regulate hypoxic breathing response. Moreover, it mentions that animal studies reveal that introduction of hypoxic stimulus in neonates produce irreversible plastic changes.

A large number of mammalian Mg2+ transporters have been hypothesized on the basis of physiological data, but few have been investigated at the molecular level. The recent identification of a number of novel proteins that mediate Mg2+ transport has enhanced our understanding of how Mg2+ is translocated across mammalian membranes. Some of these transporters have some similarity to those found in prokaryocytes and yeast cells. Human Mrs2, a mitochondrial Mg2+ channel, shares many of the properties of the bacterial CorA and yeast Alr1 proteins. The SLC41 family of mammalian Mg2+ transporters has a similarity with some regions of the bacterial MgtE transporters. The mammalian ancient conserved domain protein (ACDP) Mg2+ transporters are found in prokaryotes, suggesting an ancient origin. However, other newly identified mammalian transporters, including TRPM6/7, MagT, NIPA, MMgT, and HIP 14 families, are not represented in prokaryotic genomes, suggesting more recent development. MagT, NIPA, MMgT, and HIP14 transporters were identified by differential gene expression using microarray analysis. These proteins, which are found in many different tissues and subcellular organelles, demonstrate a diversity of structural properties and biophysical functions. The mammalian Mg2+ transporters have no obvious amino acid similarities, indicating that there are many ways to transport Mg2+ across membranes. Most of these proteins transport a number of divalent cations across membranes. Only MagT1 and NIPA2 are selective for Mg2+ Many of the identified mammalian Mg2+ transporters are associated with a number of congenital disorders encompassing a wide range of tissues, including intestine, kidney, brain, nervous system, and skin. It is anticipated that future research will identify other novel Mg2+ transporters and reveal other diseases. [ABSTRACT FROM AUTHOR]

The urine concentrating mechanism of the mammalian kidney, which can produce a urine that is substantially more concentrated than blood plasma during periods of water deprivation, is one of the enduring mysteries in traditional physiology. Owing to the complex lateral and axial relationships of tubules and vessels, in both the outer and inner medulla, the urine concentrating mechanism may only be fully understood in terms of the kidney’s three-dimensional functional architecture and its implications for preferential interactions among tubules and vessels. [ABSTRACT FROM AUTHOR]

After a complete spinal section, quadruped mammals (cats, rats, and mice) can generally regain hindlimb locomotion on a treadmill because the spinal cord below the lesion can express locomotion through a neural circuitry termed the central pattern generator (CPG). In this review, we propose that the spinal CPG also plays a crucial role in the locomotor recovery after incomplete spinal cord injury. [ABSTRACT FROM AUTHOR]

In functional genomics, the high-throughput methods such as microarrays 1) allow analysis of the relationships between genes considering them as elements of a network and 2) lead to biological interpretations thanks to Gene Ontology. But up to now it has not been possible to find relationships between the functions and the connectivity of the genes in coexpression networks. To achieve this aim, we have defined a double connectivity for each gene by the numbers of its significant negative and positive correlations with the other genes within a given biological condition, or group. Here, based on the analysis of 1,260 DNA microarrays, we show that this double connectivity clearly separates two types of genes, those with a predominantly strong negative connectivity, hub– genes, and those with a predominantly strong positive connectivity, hub+ genes. Interestingly, the hub+ genes concerned transcription factors more often than by chance and, similarly, for the hub– genes concerning miRNA predicted targets. Furthermore, a meta-analysis of GO annotations carried out on 67 groups in humans and rats shows that these two types of genes correspond to a functional biological duality. The hub– genes were mainly involved in basic functions common to all eukaryote cells, whereas the hub+ genes were mainly involved in specialized functions related to cell differentiation and communication. The separation and the biological role of these hub– and hub+ genes provide a powerful new tool for a better understanding of the control and regulation of the key genes involved in cellular differentiation and physiopathological conditions. [ABSTRACT FROM AUTHOR]

RNA interference (RNAi) has become a powerful technique for studying gene function, biological pathways, and the physiology of diseases. Typically, the RNAi response in mammalian cells is mediated by small interfering RNA (siRNA). The use of synthesized siRNA to silence gene is relatively quick and easy, but it is costly with transient effects. A short hairpin RNA (shRNA) with complementary sense and antisense sequences of a target gene separated by a loop structure results in gene silencing that is as effective as chemically synthesized siRNA with fewer limitations. However, current methods for constructing shRNA vectors require the synthesis of long oligonucleotides, which is costly and often suffers from mutation problems during synthesis. Here, we report an alternative approach to generate a shRNA expression vector with high efficacy. We utilized shorter (≤50-nucleotide) primers to generate a shRNA insert by the primer extension method. Our new approach for the construction of shRNA expression vectors dramatically reduced the possibility of mutations. Using this method, we constructed a microRNA (miRNA) library, which facilitates the expression of 254 matured miRNAs. We also performed high-throughput screening of miRNAs involved in the regulation of human Survivin promoter activity in lung A549 cells. We found that the expression of miR-192, 199a, 19a, 20a, 213, and 371 caused the activation of the Survivin promoter whereas miR-302b*, 34a, 98, 381, 463 and 471 decreased the Survivin promoter activity. [ABSTRACT FROM AUTHOR]

In mammals, the type II melanoma antigen (Mage) protein family is constituted by at least 10 closely related members that are expressed in different tissues, including the nervous system. These proteins are believed to regulate cell cycle withdrawal, neuronal differentiation, and apoptosis. However, the analysis of their specific function has been complicated by functional redundancy. In accordance with previous studies in teleosts and Drosophila, we present evidence that only one mage gene exists in genomes from protists, fungi, plants, nematodes, insects, and nonmammalian vertebrates. We have identified the chicken mage gene and cloned the cDNA encoding the chick Mage protein (CMage). CMage shares close homology with the type II Mage protein family, and, as previously shown for the type II Mage proteins Necdin and Mage-G1, it can interact with the transcription factor E2F-1. CMage is expressed in specific regions of the developing nervous system including the retinal ganglion cell layer, the ventral horn of the spinal cord, and the dorsal root ganglia, coinciding with the expression of the neurotrophin receptor p75 (p75NTR) in these regions. We show that the intracellular domain of p75NTRcan interact with both CMage and Necdin, thus preventing the binding of the latter proteins to the transcription factor E2F-1, and facilitating the proapoptotic activity of E2F-1 in N1E-115 differentiating neurons. The presence of a single mage gene in the chicken genome, together with the close functional resemblance between CMage and Necdin, makes this species ideal to further analyze signal transduction through type II Mage proteins. [ABSTRACT FROM AUTHOR]

Pulmonary surfactant, a complex mixture of lipids and proteins, lowers the surface tension in terminal air spaces and is crucial for lung function. Within an animal species, surfactant composition can be influenced by development, disease, respiratory rate, and/or body temperature. Here, we analyzed the composition of surfactant in three heterothermic mammals (dunnart, bat, squirrel), displaying different torpor patterns, to determine: 1) whether increases in surfactant cholesterol (Chol) and phospholipid (PL) saturation occur during long-term torpor in squirrels, as in bats and dunnarts; 2) whether surfactant proteins change during torpor; and 3) whether PL molecular species (molsp) composition is altered. In addition, we analyzed the molsp composition of a further nine mammals (including placental/marsupial and hetero-/homeothermic contrasts) to determine whether phylogeny or thermal behavior determines molsp composition in mammals. We discovered that like bats and dunnarts, surfactant Chol increases during torpor in squirrels. However, changes in PL saturation during torpor may not be universal. Torpor was accompanied by a decrease in surfactant protein A in dunnarts and squirrels, but not in bats, whereas surfactant protein B did not change in any species. Phosphatidylcholine (PC)16:0/16:0 is highly variable between mammals and is not the major PL in the wombat, dunnart, shrew, or Tasmanian devil. An inverse relationship exists between PC16:0/16:0 and two of the major fluidizing components, PC16:0/16:1 and PC16:0114:0. The PL molsp profile of an animal species is not determined by phylogeny or thermal behavior. We conclude that there is no single PL molsp composition that functions optimally in all mammals; rather, surfactant from each animal is unique and tailored to the biology of that animal. [ABSTRACT FROM AUTHOR]

Studies the temperature dependence of cloned mammalian and salmonid cardiac sodium/calcium exchanger isoforms. Calcium regulation of outward sodium/calcium exchange currents; Differences in current inactivation rates; Temperature dependence of the inactivation rate constant of the sodium/calcium exchange current.

Tests the hypothesis that polyamine transport system mediates agmatine transport in mammalian cells. Transport kinetics; Ion and energy dependence for agmatine and polyamine uptake in Ras/3T3 cells; Characterization of agmatine uptake.

Tests the hypothesis that the activation of P2Y but not P2X[sub4} nucleotide receptors causes elevation of intracellular calcium in mammalian osteoclasts. Calcium store depletion resulting from blockade of calcium-adenosine triphosphatase; Properties of calcium store-operated influx.

Presents information on a study which analyzed rhythmicity of cyclic guanosine monophosphate (cGMP) and cGMP-dependent kinase in suprachiasmatic nuclei in the mammalian circadian system. Methodology; Results of the study; Discussion.

Presents information on a study which investigated the adenosine monophosphate induced-regulation of the human recombinant potassium channel complex in mammalian cell lines. Methodology; Results of the study; Discussion.

Tests the hypothesis that, in mammalian muscle fibers, myosin heavy chain (MHC) isoform expression and Ca[sup 2+]- or Sr[sup 2+]-activation characteristics are only partly related. Use of chemically skinned fibers from adult rat skeletal muscles; Fiber types classified according to MHC composition; Fiber-typing methods needed to characterize the fibers.

Focuses on the development of intestinal transport function in mammals. Signals modulating the developmental changes of intestinal transport; Overview of the proliferation and differentiation of intestinal mucosa; Description of developmental patterns concerning the absorption of biological substances; Ontogeny of intestinal transport.

Examines myosin light chain replacement in the mammalian heart. Calcium-stimulated magnesium-adenosine triphosphatase activities; Mechanical analyses of skinned fiber strips from left atrial trabeculae, and left ventricular papillary muscles of non-transgenic mice; Fiber mechanics.

Clarifies the existence of extraretinal photosensitivity in mammals through the measurements of skin blood flow variation due to light irradiation. Changes in blood flow during the periods of irradiation and nonirradiation of visible light with various power densities; Wavelength dependence of the increase of blood flow.

Deals with a study on the physiology of the mammalian urinary bladder epithelium. Functions; Properties of urothelium to perform these functions; Essential requirement for normal bladder functional; Tissue culture of the urothelium.

Presents information on a study which investigates the structural identification of descending thin line (DTL)-type and ascending thin line (ATL)-type segments of single thin limbs of Henle's loop in the inner medulla. Methodology; Description of the DTL- and ATL-type segments within the rat inner medulla; Results and discussion.

Focuses on the molecular structure and function of mammalian aquaporins, a family of small membrane-spanning blood proteins. Basic features of aquaporin structure; Assembly of aquaporins in membranes as homotetramers; Potential to transport water, glycerol and larger solutes; Possible movement of ions and carbon dioxide through aquaporins.

Presents evidence linking protein kinase C to the regulation of turnover rate of rabbit intestinal Na+-glucose transporter expressed in the mammalian COS-7 cell line. Stimulation of decrease in sugar uptake by phorbol 12-myristate 13 acetate (PMA); Retention of number of cell surface transporters after PMA treatment.

Points out a resemblance between the Na+/H+ exchanger (NHE) in the red blood cells of Amphiuma tridactylum and mammals. Cloning of cDNA encoding the NHE exchanger; Sequence comparisons; Observation of Na+ dependent proton transport by Amphiuma NHE; Exhibition of drug sensitivities.

Deals with a study which characterized the intracellular calcium stores of mammalian lens cells in terms of their distribution and sensitivity to calcium releasing agents. Materials and methods of the study; Mechanism of calcium releases from intracellular stores by the adenosine diphosphate (ADP)-ribose; Spatial distribution and functional overlap of the intracellular calcium stores.