Effects of exercise and doxorubicin on acute diaphragm neuromuscular transmission failure.

Doxorubicin (DOX) is a highly effective anthracycline antibiotic used to treat a wide variety of cancers including breast cancer, leukemia and lymphoma. Unfortunately, clinical use of DOX is limited due to adverse off-target effects resulting in fatigue, respiratory muscle weakness and dyspnea. The diaphragm is the primary muscle of inspiration and respiratory insufficiency is likely the result of both muscle weakness and neural impairment. However, the contribution of neuropathology to DOX-induced respiratory muscle dysfunction is unclear. We hypothesized that diaphragm weakness following acute DOX exposure is associated with neurotoxicity and that exercise preconditioning is sufficient to improve diaphragm muscle contractility by maintaining neuromuscular integrity. Adult female Sprague-Dawley rats were randomized into four experimental groups: 1) sedentary-saline, 2) sedentary-DOX, 3) exercise-saline or 4) exercise-DOX. Endurance exercise preconditioning consisted of treadmill running for 1 h/day at 30 m/min for 10 days. Twenty-four hours after the last bout of exercise, animals were treated with DOX (20 mg/kg, I.P.) or saline (equal volume). Our results demonstrate that 48-h following DOX administration diaphragm muscle specific force is reduced in sedentary-DOX rats in response to both phrenic nerve and direct diaphragm stimulation. Importantly, endurance exercise preconditioning in DOX-treated rats attenuated the decrease in diaphragm contractile function, reduced neuromuscular transmission failure and altered phrenic nerve morphology. These changes were associated with an exercise-induced reduction in circulating biomarkers of inflammation, nerve injury and reformation. Therefore, the results are consistent with exercise preconditioning as an effective way of reducing respiratory impairment via preservation of phrenic-diaphragm neuromuscular conduction.

Loss of calpain 3 dysregulates store-operated calcium entry and its exercise response in mice.

Limb-Girdle Muscular Dystrophy 2A (LGMD2A) is caused by mutations in the CAPN3 gene encoding Calpain 3, a skeletal-muscle specific, Ca2+-dependent protease. Localization of Calpain 3 within the triad suggests it contributes to Ca2+ homeostasis. Through live-cell Ca2+ measurements, muscle mechanics, immunofluorescence, and electron microscopy (EM) in Capn3 deficient (C3KO) and wildtype (WT) mice, we determined if loss of Calpain 3 altered Store-Operated Calcium Entry (SOCE) activity. Direct Ca2+ influx measurements revealed loss of Capn3 elicits elevated resting SOCE and increased resting cytosolic Ca2+, supported by high incidence of calcium entry units (CEUs) observed by EM. C3KO and WT mice were subjected to a single bout of treadmill running to elicit SOCE. Within 1HR post-treadmill running, C3KO mice exhibited diminished force production in extensor digitorum longus muscles and a greater decay of Ca2+ transients in flexor digitorum brevis muscle fibers during repetitive stimulation. Striking evidence for impaired exercise-induced SOCE activation in C3KO mice included poor colocalization of key SOCE proteins, stromal-interacting molecule 1 (STIM1) and ORAI1, combined with disappearance of CEUs in C3KO muscles. These results demonstrate that Calpain 3 is a key regulator of SOCE in skeletal muscle and identify SOCE dysregulation as a contributing factor to LGMD2A pathology.

Targeting mitochondrial Ca2+ uptake for the treatment of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a rare adult-onset neurodegenerative disease characterized by progressive motor neuron (MN) loss, muscle denervation and paralysis. Over the past several decades, researchers have made tremendous efforts to understand the pathogenic mechanisms underpinning ALS, with much yet to be resolved. ALS is described as a non-cell autonomous condition with pathology detected in both MNs and non-neuronal cells, such as glial cells and skeletal muscle. Studies in ALS patient and animal models reveal ubiquitous abnormalities in mitochondrial structure and function, and disturbance of intracellular calcium homeostasis in various tissue types, suggesting a pivotal role of aberrant mitochondrial calcium uptake and dysfunctional calcium signalling cascades in ALS pathogenesis. Calcium signalling and mitochondrial dysfunction are intricately related to the manifestation of cell death contributing to MN loss and skeletal muscle dysfunction. In this review, we discuss the potential contribution of intracellular calcium signalling, particularly mitochondrial calcium uptake, in ALS pathogenesis. Functional consequences of excessive mitochondrial calcium uptake and possible therapeutic strategies targeting mitochondrial calcium uptake or the mitochondrial calcium uniporter, the main channel mediating mitochondrial calcium influx, are also discussed.

Comparison of Motor Relearning Program versus Bobath Approach for Prevention of Poststroke Apathy: A Randomized Controlled Trial.

BACKGROUND: Apathy is a multidimensional syndrome referring to a primary lack of motivation, frequent in survivors of stroke. And prior studies have demonstrated the negative effect of apathy on recovery from stroke. METHODS: A randomized controlled study of acute stroke patients. Four hundred and eighty-eight patients without evidence of apathy or depression at the initial visit were consecutively recruited, 258 males and 230 female. Patients were block randomized into 2 groups. Group A (n = 245) and Group B (n = 243) had physiotherapy according to Motor Relearning Program and Bobath in the first 4 weeks, respectively. The supplemental treatment did not differ in the 2 groups. Patients were assessed with Apathy Evaluation Scale-Clinical, National Institutes of Health Stroke Scale scores, Barthel Index scores, Mini-Mental State Examination scores, Hamilton Depression Scale scores, and Hamilton Anxiety Scale scores upon admission. At 1-, 3-, 6-, 9-, and 12-month follow-up after stroke, patients were assessed for diagnosis and severity of apathy using the Apathy Evaluation Scale-Clinical. RESULTS: Baseline characteristics of the subjects are age mean 65.1 (standard deviations, SD 10.9); 47.1% female; Apathy Evaluation Scale-Clinical mean 24.9 (SD 4.7); National Institutes of Health Stroke Scale mean 3.9 (SD 3.8); Barthel Index mean 87.9 (SD 8.7); Mini-Mental State Examination mean 23.3 (SD 4.5); Hamilton Depression Scale mean 17.5 (SD 6.6); and Hamilton Anxiety Scale mean 14.4 (SD 6.2). Participants in both groups had similar levels of apathy symptoms at study admission (Motor Relearning Program, mean = 24.78, SD = 4.62; Bobath, mean = 25.07, SD = 4.75). The Apathy Evaluation Scale scores of participants in both groups demonstrated to decline gradually from month 1 to month 12. Motor Learning Program participants had significantly less apathy severity compared with Bobath participants with respect to each time point. Participants given Bobath approach were 1.629 times more likely to develop poststroke apathy than patients given Motor Relearning Program over 12 months. CONCLUSIONS: Physiotherapy treatment in acute stroke rehabilitation using Motor Relearning program was significantly more effective in preventing of new onset of apathy following stroke compared with Bobath approach.

C-Reactive Protein Can Be an Early Predictor of Poststroke Apathy in Acute Ischemic Stroke Patients.

BACKGROUND: Apathy is a multidimensional syndrome referring to a primary lack of motivation that occurs frequently in survivors of stroke. Higher C-reactive protein (CRP) level was associated with higher apathy scores among Alzheimer disease cases. However, data on the relationship between CRP levels and apathy in patients with stroke are lacking. So, we hypothesized an association between CRP and poststroke apathy (PSA). METHODS: Two hundred ninety-two consecutive patients with stroke were recruited within 7 days after stroke. Apathy symptoms were assessed at baseline and at 1, 3, and 6 months after stoke using the Apathy Evaluation Scale-Clinical (AES-C). Demographic and clinical information were obtained using the National Institutes of Health Stroke Scale (NIHSS) scores, Barthel Index (BI) scores, Mini-Mental State Examination (MMSE) scores, Hamilton Depression Scale (HAMD) scores, and Hamilton Anxiety Scale (HAMA) scores. CRP was measured at baseline. The presence and the location of infarcts were evaluated using magnetic resonance imaging. RESULTS: Apathy at baseline was significantly associated with body mass index (BMI), NIHSS, BI, MMSE, HAMD, and CRP (P < .05) upon admission. PSA at 6 months was significantly associated with elevated CRP concentrations, high AES-C score, and low BI score (P < .05) upon admission. The AES-C scores peaked 3 months after stroke, but then abated over 6 months. CONCLUSIONS: CRP, BMI, MMSE, depression, and disability are closely related to apathy during the acute stage of ischemic stroke. Lower BI scores, higher CRP concentrations, and apathy in acute stroke phase increased the risk of PSA at 6 months.

Engaging in a tone-detection task differentially modulates neural activity in the auditory cortex, amygdala, and striatum.

The relationship between attention and sensory coding is an area of active investigation. Previous studies have revealed that an animal's behavioral state can play a crucial role in shaping the characteristics of neural responses in the auditory cortex (AC). However, behavioral modulation of auditory response in brain areas outside the AC is not well studied. In this study, we used the same experimental paradigm to examine the effects of attention on neural activity in multiple brain regions including the primary auditory cortex (A1), posterior auditory field (PAF), amygdala (AMY), and striatum (STR). Single-unit spike activity was recorded while cats were actively performing a tone-detection task or passively listening to the same tones. We found that tone-evoked neural responses in A1 were not significantly affected by task-engagement; however, those in PAF and AMY were enhanced, and those in STR were suppressed. The enhanced effect was associated with an improvement of accuracy of tone detection, which was estimated from the spike activity. Additionally, the firing rates of A1 and PAF neurons decreased upon motor response (licking) during the detection task. Our results suggest that attention may have different effects on auditory responsive brain areas depending on their physiological functions.

MicroRNA-107 prevents amyloid-beta induced blood-brain barrier disruption and endothelial cell dysfunction by targeting Endophilin-1.

The disruption of blood-brain barrier (BBB) and endothelial cell dysfunction, associated with the cerebrovascular deposition of the amyloid-beta (Abeta) protein, have been characterized as the key pathological characteristics in Alzheimer's disease (AD). In various biologic processes of AD, researchers have proven that mircroRNAs (miRNAs) play critical roles. However, the role and function of miRNAs in the disruption of BBB of AD still remain unclear. Here, we found that mircroRNA-107 (miR-107) is endogenously expressed in human brain microvascular endothelial cells (ECs) of BBB model, while it is significantly down-regulated in ECs pre-incubated with Abeta. Abeta significantly impairs the integrity, increases the permeability of BBB, inhibits the viability of endothelial cells (ECs), and meanwhile down-regulates the expression of tight junction proteins ZO-1, Occludin and Claudin-5. Overexpression of miR-107 largely abrogated Abeta-induced disruption of BBB and endothelial cell dysfunction. Furthermore, overexpression of miR-107 also down-regulates endophilin-1, which is involved in the regulation of BBB permeability and the expression of ZO-1, Occludin, and Claudin-5. Both bioinformatics and luciferase reporter assays demonstrated that Endophilin-1 was a direct and functional downstream target of miR-107. In conclusion, our results indicate that overexpression of miR-107 is able to prevent Abeta-induced blood-brain barrier disruption and endothelial cell dysfunction by targeting endophilin-1.

Auditory response properties of neurons in the putamen and globus pallidus of awake cats.

Several decades of research have provided evidence that the basal ganglia are closely involved in motor processes. Recent clinical, electrophysiological, behavioral data have revealed that the basal ganglia also receive afferent input from the auditory system, but the detailed auditory response characteristics have not yet reported. The present study aimed to reveal the acoustic response properties of neurons in parts of the basal ganglia. We recorded single-unit activities from the putamen (PU) and globus pallidus (GP) of awake cats passively listening to pure tones, click trains, and natural sounds. Our major findings were: 1) responses in both PU and GP neurons were elicited by pure-tone stimuli, whereas PU neurons had lower intensity thresholds, shorter response latencies, shorter excitatory duration, and larger response magnitudes than GP neurons. 2) Some GP neurons showed a suppressive response lasting throughout the stimulus period. 3) Both PU and GP did not follow periodically repeated click stimuli well, and most neurons only showed a phasic response at the stimulus onset and offset. 4) In response to natural sounds, PU also showed a stronger magnitude and shorter duration of excitatory response than GP. The selectivity for natural sounds was low in both nuclei. 5) Nonbiological environmental sounds more efficiently evoked responses in PU and GP than the vocalizations of conspecifics and other species. Our results provide insights into how acoustic signals are processed in the basal ganglia and revealed the distinction of PU and GP in sensory representation.

Behavioral modulation of neural encoding of click-trains in the primary and nonprimary auditory cortex of cats.

Neural representation of acoustic stimuli in the mammal auditory cortex (AC) has been extensively studied using anesthetized or awake nonbehaving animals. Recently, several studies have shown that active engagement in an auditory behavioral task can substantially change the neuron response properties compared with when animals were passively listening to the same sounds; however, these studies mainly investigated the effect of behavioral state on the primary auditory cortex and the reported effects were inconsistent. Here, we examined the single-unit spike activities in both the primary and nonprimary areas along the dorsal-to-ventral direction of the cat's AC, when the cat was actively discriminating click-trains at different repetition rates and when it was passively listening to the same stimuli. We found that the changes due to task engagement were heterogeneous in the primary AC; some neurons showed significant increases in driven firing rate, others showed decreases. But in the nonprimary AC, task engagement predominantly enhanced the neural responses, resulting in a substantial improvement of the neural discriminability of click-trains. Additionally, our results revealed that neural responses synchronizing to click-trains gradually decreased along the dorsal-to-ventral direction of cat AC, while nonsynchronizing responses remained less changed. The present study provides new insights into the hierarchical organization of AC along the dorsal-to-ventral direction and highlights the importance of using behavioral animals to investigate the later stages of cortical processing.

Comparison of neural responses to cat meows and human vowels in the anterior and posterior auditory field of awake cats.

For humans and animals, the ability to discriminate speech and conspecific vocalizations is an important physiological assignment of the auditory system. To reveal the underlying neural mechanism, many electrophysiological studies have investigated the neural responses of the auditory cortex to conspecific vocalizations in monkeys. The data suggest that vocalizations may be hierarchically processed along an anterior/ventral stream from the primary auditory cortex (A1) to the ventral prefrontal cortex. To date, the organization of vocalization processing has not been well investigated in the auditory cortex of other mammals. In this study, we examined the spike activities of single neurons in two early auditory cortical regions with different anteroposterior locations: anterior auditory field (AAF) and posterior auditory field (PAF) in awake cats, as the animals were passively listening to forward and backward conspecific calls (meows) and human vowels. We found that the neural response patterns in PAF were more complex and had longer latency than those in AAF. The selectivity for different vocalizations based on the mean firing rate was low in both AAF and PAF, and not significantly different between them; however, more vocalization information was transmitted when the temporal response profiles were considered, and the maximum transmitted information by PAF neurons was higher than that by AAF neurons. Discrimination accuracy based on the activities of an ensemble of PAF neurons was also better than that of AAF neurons. Our results suggest that AAF and PAF are similar with regard to which vocalizations they represent but differ in the way they represent these vocalizations, and there may be a complex processing stream between them.

[Expression of multiepitope antigenic gene of Toxoplasma gondii in transgenic tomatoes].

OBJECTIVE: To obtain the transgenic tomato plant expressed multiepitope antigenic gene of Toxoplasma gondii (Tg-MAG). METHODS: Tg-MAG under the control of E35S promoter, E35S-E81.1 chimeric promoter and E8 2.2 promoter in the plant expression vectors was transferred into the tomato cotyledons and hypocotyls via Agrobacterium-mediated T-DNA transformation. The candidate buds were obtained and elongated by kanamycin resistant screening. After further rooting selection, the selected tomato lines were trained and transplanted to soil and grown in a greenhouse till blossoming and bearing fruit. The transgenic tomato plants were identified using PCR, RT-PCR and Western blotting. RESULTS: Tg-MAG was introduced into the transgenic plant genomic DNA and transcribed correctly at expected size 360 bp identified by PCR and RT-PCR. Western blotting results indicated that the recombinant Tg-MAG from 8 transgenic lines, expressed in tomato fruits at expected size 11900, 2 of them showed strong reaction band with HRP labeled McAb K7H3 against the major surface antigen 1( SAG1) of T. gondii tachyzoites. CONCLUSION: Tg-MAG transgenic tomato lines were obtained. Tg-MAG recombinant protein with good immune activity was expressed successfully in transgenic tomato fruits.