Comparison of the antioxidant status in tracheal and bronchoalveolar epithelial lining fluids in recurrent airway obstruction.

REASONS FOR PERFORMING STUDY: Following a period of airway inflammation the clearance of inflammatory cells along the mucociliary escalator may impose a considerable oxidant load on the trachea. OBJECTIVES: To determine the degree of oxidative stress in tracheal epithelial lining fluid (ELF) in comparison to that present in peripheral airways after an acute exposure to organic dust. METHODS: Tracheal wash fluid and bronchoalveolar lavage fluid (BALF) were collected for cytology and antioxidant analyses from 6 recurrent airway obstruction (RAO)-affected horses and 6 healthy control horses before and after stabling on straw bedding for 24 h. RESULTS: In RAO-affected horses, organic dust exposure resulted in a significant decrease in ascorbic acid concentration in tracheal ELF (P<0.0001), which was greater than the decrease in bronchoalveolar ELF (P = 0.0003). The percentage decrease in tracheal ELF ascorbic acid correlated with the percentage decrease in bronchoalveolar ELF ascorbic acid (r = 0.76; P = 0.004) following exposure. CONCLUSIONS: Acute organic dust exposure results in significant antioxidant depletion in the trachea, which may reflect inflammation and oxidative processes in peripheral airways. POTENTIAL RELEVANCE: Further work is required to evaluate the role of ascorbic acid depletion in the pathogenesis of RAO.

The salivary gland and salivary enzymes of the giant waterbugs (Heteroptera; Belostomatidae).

The giant waterbugs are predators that utilize extra-oral digestion and are known to capture a wide variety of prey. Herein we describe the differences in salivary enzyme composition between large and small species of giant waterbug (Lethocerus uhleri, Lethocerinae and Belostoma lutarium, Belostomatinae, respectively). The saliva of L. uhleri contains 3 proteolytic enzymes and no amylase, while the salivary gland of B. lutarium produces 2 proteolytic enzymes and amylase. This fundamental difference in salivary enzyme composition correlates with the difference in diet preference between the Lethocerinae and Belostomatinae. Furthermore, we describe the ultrastructure of the salivary gland complex of B. lutarium and present data on the division of labor with respect to compartmentalization of enzyme production. Proteolytic enzymes are produced in the accessory salivary gland and amylase is produced in the main salivary gland lobe. This is the first reported evidence of protease production in the accessory salivary gland in the Heteroptera.

Bulbus arteriosus of the bivalve mollusc Mercenaria mercenaria: Morphology and pharmacology.

We examined the morphology and pharmacology of the bulbus arteriosus of the marine bivalve mollusc Mercenaria mercenaria. The bulbus arteriosus is an organ of unknown function associated with the posterior aorta and the ventricle. It is composed of connective tissue interspersed with muscle bundles. In contrast to the lumen of the ventricle, which has an extensive trabecular network, the lumen of the bulbus arteriosus has no trabeculae. No valve is present in the posterior aorta connecting the lumina of the ventricle and the bulbus arteriosus. Scattered neuronal profiles are present in the wall and the axonal processes contain vesicles that may contain neurosecretory products. We did not, however, find synapses or evidence of vesicular release into the lumen of the bulbus arteriosus. The bulbus arteriosus contains acetylcholine, 5-hydroxytryptamine (5HT), and the molluscan neuropeptides FMRFamide (phenyl-alanyl-methionyl-arginyl-phenylalaninylamide) and FLRFamide (phenylalanyl-leucinyl-arginyl-phenylalanylamide). The isolated bulbus arteriosus contracts tonically in response to mechanical stress and exposure to 5HT or FMRFamide, while acetylcholine relaxes it. We surmise that the bulbus arteriosus probably functions as a mechanism for regulating the relative amounts of hemolymph pumped into the anterior and posterior aortae by the ventricle and that the bulbus arteriosus may be a neurohemal site.

Hyperosmotic volume regulation in the gills of the ribbed mussel, Geukensia demissa: rapid accumulation of betaine and alanine.

The content of betaine and alanine in gills of the ribbed mussel Geukensia demissa increases rapidly following transfer of the tissues from 250 to 1000 mOsm seawater (SW). Increases in alanine, proline and glycine account for most of the increase in the amino acid pool. The betaine content increases from 45 to 150 &mgr;mol/g dry weight within 12 h. Transfer of isolated gills from 250 to 1000 mOsm SW results in a temporary cessation of all ciliary activity. Within 20-40 min following transfer, ciliary activity has recovered. Recovery of ciliary activity precedes recovery of tissue hydration. The uric acid content of gills is unchanged by exposure to hyperosmotic media, suggesting that uric acid is not a store of nitrogen for alanine synthesis from pyruvate. In other organisms, the accumulation of betaine in response to hyperosmotic stress is a slow (days to weeks) process that probably involves changes in gene expression. The rapid, large increases in betaine reported here suggest that gene expression is not a factor in volume recovery by euryhaline bivalve tissues exposed to acute hyperosmotic stress.

Comparative aspects of cellular-volume regulation in cardiomyocytes.

All cells possess mechanisms that are responsible for the maintenance of cellular volume under isosmotic conditions. In addition, many cells are able to adjust cellular volume when incubated in hypo- or hyperosmotic media. Much of the work on cellular-volume regulation has been done on epithelial cells, blood cells, or lines of cultured cells; cardiac muscle has received comparatively little attention. It seems probable that some aspects of cellular-volume regulation in cardiomyocytes vary from those present in other cell types because of the mechanisms associated with the excitability and contractility of cardiac muscle. For example, in myogenic hearts, the role of membrane ion channels in pacemaker potentials complicates models that implicate ion channels as mechanisms for volume regulation. Similarly, models for the initiation and control of volume regulation that rely on changes in cytosolic Ca2+ levels may not be applicable to cardiac muscle, where each action potential and contraction involves the release of Ca2+ from internal stores and a significant influx of Ca2+ across the plasma membrane. A review of the available data on volume regulation in cardiac muscle from a variety of invertebrate and vertebrate species suggests that many features of the current models proposed for the initiation and control of cellular-volume regulation are not compatible with the physiology of cardiac muscle. There are large gaps in our knowledge about volume regulation in cardiac muscle, and further investigation is clearly necessary to enhance our understanding of this aspect of cardiomyocyte physiology.

The effects of FMRFamide, 5-hydroxytryptamine and phorbol esters on the heart of the mussel Geukensia demissa.

The ventricle of the mussel Geukensia demissa is inhibited by 5-hydroxytryptamine and excited by the molluscan neuropeptide FMRFamide. Supra-threshold doses of amide result in marked positive chronotropy and inotropy within 5-15 s. 5-Hydroxytryptamine at 10(-8) M produces diastolic arrest within 10 s. A 1-min exposure to FMRFamide (5 x 10(-8) M) results in a small increase in the cytoplasmic levels of adenosine 3',5'-cyclic monophosphate; shorter or longer exposures have no effect. The cAMP content of ventricles incubated in 5 x 10(-8) M 5-hydroxytryptamine for 1 min decreases by 2.3 pmol/mg protein; longer or shorter incubations have no effect. Treatment with forskolin results in 3- or 4-fold increases in adenosine 3',5'-cyclic monophosphate, but forskolin has no effect on the mechanical activity of the ventricle. The levels of inositol monophosphate, inositol 1,4-diphosphate, and inositol 1,4,5-triphosphate in tissues exposed to 5-hydroxytryptamine are not different from levels in control tissues. FMRFamide decreases the levels of these phosphoinositides by 50% or more. Lower concentrations of phorbol 12,13-diacetate (10(-8) to 10(-7) M) and phorbol 12-myristate,13-acetate (10(-6) M) cause positive chronotropy in the isolated ventricle; higher concentrations induce systolic arrest. These results suggest that the effects of 5HT on the ventricle are not mediated by adenosine 3',5'-cyclic monophosphate or inositol 1,4,5-triphosphate. The effects of FMRFamide may involve a decrease in inositol 1,4,5-triphosphate. The effects of amide may involve a decrease in inositol 1,4,5-triphosphate. The response of the ventricles to phorbol esters suggest that protein kinase C may be involved in the regulation of cardiac contractility.

Potentiation of Hypoosmotic Cellular Volume Regulation in the Quahog, Mercenaria mercenaria, by 5-hydroxytryptamine, FMRFamide, and Phorbol Esters.

Ventricles isolated from clams (Mercenaria mercenaria) that had been acclimated to 1000 mOsm seawater (SW) release amino acids when incubated in 500 mOsm SW. Taurine, glycine, and alanine account for nearly all of the released amino acids, and total about 37 µmol/g dry tissue weight during a 2-h incubation. The release of amino acids is increased to 69 µmol/g by the addition of 10-6 M 5-hydroxytryptamine (5HT) to the hypoosmotic SW, and to 83 µmol/g by the addition of 10-6 M FMRFamide to the medium. The potentiation of the release by 5HT is blocked by methysergide. The amino acid release is increased by two phorbol esters--phorbol 12,13-diacetate and phorbol 12-acetate, 13-myristate--to 97 and 83 µmol/g, respectively. Forskolin and other cyclic 3',5' adenosine monophosphate agonists have no effect on the release of amino acids in hypoosmotic SW. Phorbol esters, 5HT, and FMRFamide have no effect on the release of amino acids from ventricles incubated in 1000 mOsm SW. Ventricles, first isolated from clams acclimated to 1000 mOsm SW, and then transferred to 500 mOsm SW, increase in wet weight by 20-25%. The increase is maintained for 30 min, and the tissues return their original weight in the ensuing 30 min. The addition of 5HT, FMRFamide, or phorbol esters to the hypoosmotic SW decreases the time necessary for the tissues to return to pre-transfer weights. These results implicate protein kinase C in the responses of bivalve tissues to hypoosmotic media, and suggest that these responses may be modified by neuronal or neurohumoral control.