Nitric oxide myotoxicity is age related.

Nitric oxide (NO) is generated under normal conditions in skeletal muscle and acts as a messenger that influences contractility, blood flow, and glucose metabolism. Excess NO generation may occur in pathological states, in particular inflammatory conditions. We demonstrate that incubation of rat extensor digitorum longus muscle with the NO donor, S-nitrosocysteine, leads to release of creatine kinase, a marker of muscle injury after a delay of 90 min. Muscle of old animals was more sensitive to the NO donor. Light microscopic analysis does not show abnormalities, with the exception of an increase in interfiber distance. Histological staining identified no pathological elevations of calcium. The study demonstrates the direct toxicity of NO to skeletal muscle, and that muscle of older animals is differentially susceptible to NO toxicity.

Nitric oxide synthase in aging rat skeletal muscle.

The neuronal isoform of nitric oxide synthase (NOS) is expressed at high concentrations in skeletal muscle, and NO influences muscle contractility, glucose utilization, and free radical damage or protection. NOS activity and expression was evaluated in extensor digitorum longus (EDL), soleus, and diaphragm of 8 and 24 month old Fisher 344 rats. In 8-month-old animals, NOS activity was highest in EDL, which contained the highest percentage of NOS containing fibers, and was lowest in soleus. NOS activity and percentage of NOS containing fibers was significantly reduced in all muscle groups with age. To determine if NOS reduction correlated with free radical injury the level of lipid peroxidation, as measured by malonaldehyde equivalents, was determined. With age lipid peroxidation increased in EDL, was reduced in diaphragm, and showed a non-significant change in soleus. Therefore, a straightforward reduction of NOS activity does not correlate with lipid peroxidation. The reduction of NOS with age in skeletal muscle may be most significant for muscle metabolism and force production and be of limited significance for free radical metabolism.

Effects of diazinon on the gills of bluegill sunfish Lepomis macrochirus.

Damaging effects of diazinon, an organophosphorus insecticide, on the gills of bluegills, Lepomis macrochirus, are many. Adult bluegills were exposed to 15, 30, 45, 60, and 75 micrograms/L of diazinon and their gills were microscopically examined. Compared to the gill structure of the control fish, all the diazinon concentrations caused various types of changes, such as lifting of the epithelial layer, hyperplasia and necrosis, shortening of the lamellae and frequent epithelial rupture, lamellar fusion, severe hyperplasia and mucous cells hypertrophy, extensive fusion, and clavate lamellae. Some of the structural changes may serve as a defense mechanism in protecting the fish from the diazinon-contaminated water by increasing the diffusion distance. Increase in the diffusion distance and mucus secretion can adversely affect the respiration of the fish, ultimately leading to its death.

Brain acetylcholinesterase activity and optomotor behavior in bluegills, Lepomis macrochirus exposed to different concentrations of diazinon.

Organophosphorus pesticides inhibit acetylcholinesterase (AChE) activity to varying degrees in different species of fish. Estimation of AChE activity is a useful tool for establishing the degree of pollution caused by these pesticides. Behavioral changes are very good and sensitive indicators of any chemical pollution. Changes in the optomotor behavior are easily quantifiable. Therefore estimation of AChE activity and changes in the optomotor response are taken as criteria to study the effects of different concentrations (15, 30, 45, 60 and 75 micrograms/l) of diazinon, an organophosphorus compound, in bluegill Sunfish, Lepomis macrochirus. The fish were exposed for a period of 24 hours. In this study, AChE activity showed a significant decline above an exposure concentration of 45 micrograms/l exposure concentration. In the case of optomotor behavior, a decline in the scores of the "following" responses of the fish was noticed from an exposure concentration of 30 micrograms/l. It seems that a behavioral bioassay is more sensitive than other types of testing.

Hypoglossal and phrenic motoneuron responses to serotonergic active agents in rats.

5-Hydroxytryptamine (serotonin, 5-HT) affects upper airway and chest wall inspiratory muscle control. The purpose of this study was to investigate the relative interaction of serotonergic agents on these two muscle groups. We measured the responses of the hypoglossal and phrenic nerves to the systemic administration of serotonergic-active agents and determined the receptor types through which these agents act in anesthetized, vagotomized, paralyzed and artificially ventilated rats. The serotonin precursor, L-5-hydroxytryptophan (L-5-HTP) produced equivalent stimulation of phasic inspiratory activity of the hypoglossal and phrenic nerves. General serotonin antagonists produced significant and equivalent diminution of both motoneuron pools. Specific 5-HT1A stimulation and 5-HT1C/2 antagonism enhanced ventilatory activity. We conclude: (1) a baseline level of serotonergic input to hypoglossal and phrenic motoneuron pools was present, (2) different 5-HT receptors had different effects on ventilatory neural activity, and (3) hypoglossal and phrenic motoneuron pools responded similarly to the serotonergic agents given.

Nitric oxide synthase expression and effects of nitric oxide modulation on contractility of rat extraocular muscle.

Extraocular muscles (EOMs) are specialized skeletal muscles that are constantly active, generate low levels of force for cross sectional area, have rapid contractile speeds, and are highly fatigue resistant. The neuronal isoform of nitric oxide synthase (nNOS) is concentrated at the sarcolemma of fast-twitch muscles fibers, and nitric oxide (NO) modulates contractility. This study evaluated nNOS expression in EOM and the effect of NO modulation on lateral rectus muscle's contractility. nNOS activity was highest in EOM compared with diaphragm, extensor digitorum longus, and soleus. Neuronal NOS was concentrated to the sarcolemma of orbital and global singly innervated fibers, but not evident in the multi-innervated fibers. The NG-nitro-L-arginine methyl ester (L-NAME, a NOS inhibitor), increased submaximal tetanic and peak twitch forces. The NO donors S-nitroso-N-acetylcysteine (SNAC) and spermineNONOate reduced submaximal tetanic and peak twitch forces. The effect of NO on the contractile force of lateral rectus muscle is greater than previously observed on other skeletal muscle. NO appears more important in modulating contraction of EOM compared with other skeletal muscles, which could be important for the EOM's specialized role in generation of eye movements.

The carotid body in the motorneuron response to protriptyline.

Protriptyline (PRT) has been shown to preferentially stimulate upper airway inspiratory motorneurons relative to phrenic activity in hyperoxic hypercapnia in the decerebrate cat via a carotid body-independent mechanism. Since previous studies indicated that carotid body stimulation results in preferential activation of upper airway respiratory muscles during both hypercapnia and hypoxemia, we hypothesized that if PRT preferentially stimulated upper airway motorneurons, the mechanism of action might involve the carotid body. We investigated the effect of PRT on carotid body function by comparing the electrical activity of the hypoglossal (HYP) with that of the phrenic (PHR) nerve in carotid sinus nerve intact (CSNI) and CSN-sectioned (CSNX) anesthetized rats, before and after PRT (0.5 mg/kg i.v.), during 100% O2, 15% O2 (N2 balance), and 4% CO2 (O2 balance) administration. The moving time average (MTA) peak inspiratory electroneurogram activities of both the HYP and PHR nerves increased an equivalent amount after PRT injection during hyperoxia, in both CSNI and CSNX rats. During hypoxia, the HYP activity increased significantly more than the PHR activity only in CSNI rats after PRT injection. During hyperoxic hypercapnia, HYP MTAs increased a similar amount in the CSNI and CSNX rats. We conclude that the HYP and PHR respiratory motorneuron pool responses to PRT depend on the blood gas status at the time of drug administration.

Human fetal membranes release a Ca++ channel inhibitor.

OBJECTIVE: Our purpose was to test the hypothesis that an inhibitor of uterine contractions released by human fetal membranes acts on the dihydropyridine site of the myometrial voltage-dependent Ca++ L channel. STUDY DESIGN: Initial experiments established the time course of release of the inhibitor from term, fetal membranes. Both a competitive binding assay and a uterine contraction bioassay were used to detect the inhibitor. After optimal time of release of inhibitor was determined, a dose-response experiment was performed with the competitive binding assay. To determine the source of the inhibitor, membranes are separated into component layers to generate inhibitor, and the competitive binding assay was used to measure the inhibitor. RESULTS: An inhibitor released from fetal membranes competes with 3H-isradipine at the Ca++ L channel dihydropyridine binding site. There is a time-dependent release of the inhibitor from membranes, which is maximal at 20 minutes (P < or = .05, n = 4). A dose effect of the inhibitor is present because greater amounts of inhibitor produce greater competition at the dihydropyridine site (P < or = .005, n = 3). The data are consistent with 1-site binding. Inhibition is restricted to the chorion (64% specific inhibition) and decidua (52% specific inhibition) with little competition seen in amnion alone (4% specific inhibition) (P < or = .03, n = 3). CONCLUSIONS: These studies support the hypothesis that human chorion/decidua releases an inhibitor of uterine contractions that acts specifically at the dihydropyridine site of the myometrial Ca++ L channel.

Extraocular muscle is defined by a fundamentally distinct gene expression profile.

Skeletal muscle fibers are defined by patterned covariation of key traits that determine contractile and metabolic characteristics. Although the functional properties of most skeletal muscles result from their proportional content of a few conserved muscle fiber types, some, typically craniofacial, muscles exhibit fiber types that appear to lie outside the common phenotypic range. We analyzed gene expression profiles of three putative muscle classes, limb, masticatory, and extraocular muscle (EOM), in adult mice by high-density oligonucleotide arrays. Pairwise comparisons using conservative acceptance criteria identified expression differences in 287 genes between EOM and limb and/or masticatory muscles. Use of significance analysis of microarrays methodology identified up to 400 genes as having an EOM-specific expression pattern. Genes differentially expressed in EOM reflect key aspects of muscle biology, including transcriptional regulation, sarcomeric organization, excitation-contraction coupling, intermediary metabolism, and immune response. These patterned differences in gene expression define EOM as a distinct muscle class and may explain the unique response of these muscles in neuromuscular diseases.