Neuromuscular defects and breathing disorders in a new mouse model of spinal muscular atrophy.

Spinal muscular atrophy (SMA) is caused by insufficient levels of the survival motor neuron (SMN) protein leading to muscle paralysis and respiratory failure. In mouse, introducing the human SMN2 gene partially rescues Smn(-)(/)(-) embryonic lethality. However current models were either too severe or nearly unaffected precluding convenient drug testing for SMA. We report here new SMN2;Smn(-/-) lines carrying one to four copies of the human SMN2 gene. Mice carrying three SMN2 copies exhibited an intermediate phenotype with delayed appearance of motor defects and developmental breathing disorders reminiscent of those found in severe SMA patients. Although normal at birth, at 7 days of age respiratory rate was decreased and apnea frequency was increased in SMA mice in parallel with the appearance of neuromuscular junction defects in the diaphragm. With median survival of 15 days and postnatal onset of neurodegeneration, these mice could be an important tool for evaluating new therapeutics.

[VOLTAGE-GATED CALCIUM CHANNELS: CLASSIFICATION AND PHARMACOLOGICAL PROPERTIES (PART I).]

Calcium influx though voltage-gated calcium channels mediate a huge amount of physiological events and cellular responses. Numerous scientific reports indicate that calcium channels are involved in synaptic transmission, neurotransmitter release, regulation of gene expression, cellular membrane voltage oscillations, pacemaker activity, secretion of specific substances from nerve and secretory cells, morphological differentiation, activation of calcium-dependent enzymes, etc. This review represents the modern classification, molecular structure, physiological and pharmacological properties of voltage-gated calcium channels expressed in mammalian cells.

Adenosine and neurotrauma: therapeutic perspectives.

Cerebral ischemia studies demonstrating that stimulation of adenosine A1 receptors by either endogenously released adenosine or the administration of selective receptor agonists causes significant reductions in the morbidity and mortality associated with focal or global brain ischemias have triggered interest in the potential of purinergic therapies for the treatment of traumatic injuries to the brain and spinal cord. Preliminary findings indicate that activation of A1 adenosine receptors can ameliorate trauma-induced death of central neurons. Other avenues of approach include the administration of agents which elevate local concentrations of adenosine at injury sites by inhibiting its metabolism to inosine by adenosine deaminase, rephosphorylation to adenosine triphosphate by adenosine kinase; or re-uptake into adjacent cells. Amplification of the levels of endogenously released adenosine in such a 'site and event specific' fashion has the advantage of largely restricting the effect of such inhibitors to areas of injury-induced adenosine release. Another approach involving purinergic therapy has been applied to the problem of respiratory paralysis following high spinal cord injuries. In this instance, the adenosine antagonist theophylline has been used to enhance residual synaptic drive to spinal respiratory neurons by blocking adenosine A1 receptors. Theophylline induced, and maintained, hemidiaphragmatic recovery for prolonged periods after C2 spinal cord hemisection in rats and may prove to be beneficial in assisting respiration in spinal cord injury patients.

Restoration of the functions of the respiratory center in rats after total cold paralysis.

White rats were cooled in water until respiratory arrest, which appeared at a brain temperature of 17.62 +/- 0.16 degrees C; animals were then removed from the water to a room at a temperature of 19-20 degrees C. Experimental animals were given 1 ml of 0.6% EDTA, which binds calcium ions, 1 min after the onset of respiratory arrest; controls received 1 ml of physiological saline. Electrical heaters were used to warm the medulla oblongata; respiratory recovery occurred at 6.0 +/- 0.3 min from the onset of respiratory arrest, when the medulla oblongata temperature was 18.62 +/- 0.15 degrees C. In controls, heating of the medulla oblongata to 18.84 +/- 0.17 degrees C did not restore respiration. EDTA bound calcium ions, facilitating transport of the excess quantities of these ions from the cell cytosol to the intercellular medium. Decreases in the cytosol calcium ion concentration to normal restored the normal regulation of metabolism in cells.

[Role of nitric oxide in diaphragmatic dysfunction genesis during sepsis in rats]. / Rôle du monoxyde d'azote (NO) dans la genèse de la dysfonction diaphragmatique au cours du sepsis chez le rat.

Nitric oxide (NO) is a vasodilator agent that is cytotoxic and negatively inotropic in the heart. More recently, it has been shown that during sepsis there is a high amount of NO production by a NO synthase (NOS) that is inducible by cytokines. The aim of this study was to investigate the role of NO in the genesis of diaphragmatic dysfunction during sepsis. Rats were inoculated i.p. injection with 10 mg/kg of Escherichia coil endotoxin (E animals) or saline (C animals). Six hours after endotoxin or saline inoculation, diaphragmatic force and muscularc GMP (Cyclic guanosine monophosphate) were assessed by in vitro force frequency curves and ELISA method, respectively. As compared to C animals, E animals showed a significant decrease in diaphragmatic force for all the frequencies of stimulation (p < 0.01). This reduction was associated with a significant increase in muscular cGMP. Inhibition of NO synthesis in E animals with either dexamethasone (4 mg/kg IV, 45 min before endotoxin or saline) or NG-monomethyl-L-arginine (L-NMMA, 8 mg/kg IV, 90 min after endotoxin or saline) prevented the effects of endotoxin. However, no modification was seen with NG-monomethyl-D-arginine (D-NMMA), a molecule which does not inhibit NO synthesis. Administration of dexamethasone or L-NMMA in C animals did not induce any significant change in diaphragmatic force, and cGMP ratio. We conclude that NO has a contributive role in diaphragmatic dysfunction during Escherichia coli induced sepsis in rats.

ERS Meeting Report. Metabolic aspects of obstructive sleep apnoea syndrome.

Insulin resistance is often associated with obstructive sleep apnoea syndrome (OSAS) and could contribute to cardiovascular risk in OSAS. Sleep loss and intermittent hypoxia could contribute to the pathogenesis of the metabolic alterations associated with obesity, a common feature of OSAS. The biology of the adipocyte is being increasingly studied, and it has been found that hypoxia negatively affects adipocyte function. In November 2007, the European Respiratory Society and two EU COST Actions (Cardiovascular risk in OSAS (B26) and Adipose tissue and the metabolic syndrome (BM0602), held a Research Seminar in Düsseldorf, Germany, to discuss the following: 1) the effects of hypoxia on glucose metabolism and adipocyte function; 2) the role of inflammatory activation in OSAS and obesity; 3) the alarming rates of obesity and OSAS in children; 4) the harmful effects of the metabolic syndrome in OSAS; 5) the effects of OSAS treatment on metabolic variables; and 6) the relationship between daytime sleepiness and hormonal and inflammatory responses. Insulin resistance in skeletal muscle, the role of the endocannabinoid system and novel pharmacological approaches to treat insulin resistance were also discussed. As obesity and hypoxia could be the basic links between OSAS and adipocyte dysfunction, further research is needed to translate these new data into clinical practice.

Sustained oxygenation without ventilation in paralyzed pigs with high-flow tracheal oxygen.

OBJECTIVES: It is generally assumed that ventilation is necessary for oxygenation. This study tested if paralyzed animals without respirations can maintain arterial oxygenation when administered high-flow oxygen delivered by a catheter in the trachea. DESIGN: Prospective observational study. SETTING: University research laboratory. PARTICIPANTS: 3 anesthetized/paralyzed swine weighing 29.5 +/- 4.2 kg. INTERVENTIONS/OBSERVATIONS: Pigs were intubated, anesthetized with intravenous tiletamine and a pentobarbital drip. A femoral arterial line was placed to record arterial blood gases and vital signs every 5 minutes. Respiratory paralysis was obtained with vecuronium 150 microg/kg and repeated at any sign of movement. A catheter was placed in the trachea to deliver oxygen at 15 L/min. Outflow gas from the endotracheal tube was analyzed for O2 and CO2. O2 was discontinued at 75 minutes. The institutional animal care and use committee approved the protocol. RESULTS: All pigs survived to 75 minutes. PaO2 was more than 100 mm Hg throughout the study period. Mean PaCO2 was 37.4 +/- 2.8 mm Hg at baseline, 146 +/- 59 at 30 minutes, then rose above 200 mm Hg in all pigs by 45 minutes. Mean arterial pH fell from 7.47 +/- 0.04 at onset to 6.75 +/- 0.06 at 75 minutes. When oxygen was terminated at 75 minutes, PaO2 fell to 16.5 +/- 7.6 mm Hg within 5 minutes, and all pigs were sacrificed within 10 minutes. For outflow gas, O2 was more than 98% and expired CO2 less than 1% throughout the study period. CONCLUSIONS: Paralyzed, unventilated pigs receiving high-flow oxygen via a tracheal catheter remained alive after 75 minutes, although a profound respiratory acidosis developed.

Hyperventilation in the newborn piglet does not increase whole body oxygen consumption as seen in mature animals.

Hyperventilation has been shown to cause increased whole body oxygen consumption (VO2) and lactic acid production in human and animal mature subjects, but limited data are available in neonates. We investigated the effect of hypocarbic and normocarbic hyperventilation during normoxia and hypoxia (fractional inspired oxygen concentration = 0.14) upon the VO2 in anesthetized and paralyzed piglets. Systemic arterial, pulmonary arterial, and left and right atrial pressures as well as cardiac output and body temperature were continuously recorded. Hypocarbic hyperventilation (PaCO2 = 19 +/- 1 mm Hg; pH = 7.58 +/- 0.02) was associated with a significant decrease in systemic and pulmonary arterial pressures and cardiac output. These measurements returned to values similar to the initial normoventilation ones when PaCO2 was increased by adding CO2 to the inspired gas, whereas hyperventilation was continued. Neither hyperventilation alone nor in combination with hypoxia induced any significant change in VO2. We conclude that in the newborn pig, unlike what has been reported in mature subjects, cellular metabolic function is unaffected by hyperventilation as evidenced by the unchanged VO2.

[Restoration of the respiratory center functions in rats after its complete cold-induced paralysis ]. / Vosstanovlenie funktsii dykhatel'nogo tsentra u krys posle ego polnogo kholodovogo paralicha.

White rats were cooled until cessation of their breathing (about 17.6 degrees C) and then transferred to a 19-20 degrees C room. The breathless rats were administered EDTA (experimental group) and saline (control groups). Within 6 min., warming of the brain stem to 18.6 degrees C caused restoration of breathing in experimental rats whereas the control rats remained breathless. The EDTA binds calcium ions in the cells' cytosol and thus restores the normal regulation of metabolism in the brain cells.

Respiratory insufficiency in desminopathy patients caused by introduction of proline residues in desmin c-terminal alpha-helical segment.

Mutations in desmin gene have been identified in patients with cardiac and skeletal myopathy characterized by intracytoplasmic accumulation of desmin-reactive deposits and electron-dense granular aggregates. We characterized two new desminopathy families with unusual features of adult-onset, slowly progressive, diffuse skeletal myopathy and respiratory insufficiency. Progressive reduction of respiratory muscle strength became clinically detectable between the 3rd and the 8th years of illness and led to recurrent chest infections and death in one of the patients. Novel mutations, A357P and L370P, predicted to introduce proline residue into a highly conserved alpha-helical region of desmin, were identified. Proline is known to disrupt the alpha-helix. In addition, the A357P mutation distorts a unique stutter sequence that is considered to be critically important for proper filament assembly. Functional assessment in two cell-lines, one of which does and the other of which does not constitutively produce type III intermediate filaments, demonstrated the inability of mutant desmin carrying either the A357P or the L370P mutation to polymerize and form an intracellular filamentous network. The results of this study indicate that respiratory insufficiency is an intrinsic feature of disease associated with specific desmin mutations; in some patients, respiratory weakness may present as a dominant clinical manifestation and a major cause of disability and death.