Follicle stimulating hormone (FSH) acting on Sertoli cells of the testis plays important roles during reproductive development. FSH-R knockout (FORKO) mice provide a model to examine alterations in testicular and epididymal structure and function in its absence. Examination of the FORKO testis revealed a gross alteration of Sertoli cell structure indicative of a fluid imbalance. Functional parameters, such as ABP secretion were also significantly reduced in FORKO testis. Morphometry revealed quantitative reductions in seminiferous tubule size. The epididymal epithelium, appeared abnormal and morphometry revealed that epididymal tubule size was reduced in the knockout. Computer Assisted Sperm Analysis on sperm from the cauda epididymidis revealed significant alterations in parameters corresponding to sperm motility as well as sperm counts. These data suggest an important role for the FSH receptor on Sertoli cell structure and functions and on epididymal epithelial size and functions in relation to sperm motility.

Thalassemia is a blood disorder requiring lifelong transfusions for survival. Erythrocytes accumulate toxic iron at their membranes, triggering an oxidative cascade that leads to their premature destruction. We hypothesized that removing this proximate iron compartment as a primary treatment using novel iron chelators, could prevent hastened red cell removal and clinically alleviate the need for transfusion. Novel, highly cell permeable iron chelators, pyridoxal isonicotinoyl hydrazone (PIH) and pyridoxal ortho-chlorobenzoyl hydrazone (o-108) were compared to the present mainstay, desferrioxamine (DFO) and deferiprone (L1), in vitro and in vivo . Treatment of human model beta-thalassemic erythrocytes with chelators resulted in significant depletion of membrane-associated iron and reduced oxidative stress as indicated by a decrease in methemoglobin levels. When administered to beta-thalassemic mice, iron chelators mobilized erythrocyte membrane iron, reduced cellular oxidation, and prolonged erythrocyte survival. Consistently, these mice showed improved hematological abnormalities. A beneficial effect as early as the erythroid precursor stage was also determined by normalized proportions of mature versus immature reticulocytes. Remarkably, all four chelators reduced iron accumulation in target organs. Most importantly, o-108 revealed superior activity, decreasing iron in liver and spleen by ~5-fold and ~2-fold, respectively, compared to DFO. Our study demonstrates that iron chelators ameliorate thalassemia in a human and murine model, and validates their primary use as an alternative to transfusion therapy.

In glomerular epithelial cells (GEC), the actin cytoskeleton is a key determinant of cell morphology and functions, including permselectivity. Complement C5b-9 induces sublytic GEC injury associated with GEC morphological changes and proteinuria. This study addressed the role of Rho GTPases in complement-mediated GEC injury. We demonstrated that the amount of active RhoA increased; while the amount of active Rac1 and Cdc42 were decreased in C5b-9 mediated sublytic GEC injury both in vitro and in glomeruli from rats with PHN in vivo. Complement mediated inactivation of p190RhoGAP may contribute to complement-induced RhoA activation. Overexpression of constitutively active or dominant negative mutants of RhoA, Rac1 and Cdc42 distinctly altered GEC morphology and F-actin pattern. Complement caused changes in GEC actin cytoskeleton, at least in part mediated by a downstream kinase of RhoA--Rho kinase (ROCK). Activation of RhoA exacerbated complement-mediated cytotoxicity in GEC, while inhibition of ROCK attenuated it.

Airway hyperresponsiveness (AHR) is a key feature of asthma, characterized by exaggerated rate and extent of shortening of airway smooth muscle. Two isoforms of the smooth muscle myosin differ by the presence [(+)insert] or absence [(-)insert] of a 7 amino acid insert. The (+)insert exhibits a 2-fold greater ATPase activity and velocity of actin filament propulsion in the in vitro motility assay. The expression of these isoforms and other contractile proteins was quantified in the trachea of the Fisher and Lewis rat model of innate AHR. We found 95% greater mRNA and 45% greater protein expression of the (+)insert isoform in the trachea of the hyperresponsive Fisher animal (p<0.01), but no difference in other contractile proteins. A greater extent of myosin phosphorylation was also observed (55.1+/- d6.4 vs. 41.4+/-d6.1, p<0.01). These results suggest that in addition to greater myosin activation, an increased expression of the (+)insert isoform contribute to AHR.

No description available.

The present investigations were aimed at providing an empirical basis for implicating altered prostaglandin (PG) and thromboxane (TX) biosynthesis in the aberrant heme metabolism and erythrocyte membrane properties which have been found to exist in hereditary muscular dystrophy and ataxia. / Picomolar concentrations of PGE(,1) and PGE(,2) were shown to exert opposing, biphasic effects on osmotic fragility, the hemolytic response to ouabain (without any direct effect on cation-activated adenosine triphosphatases), and echinocyte transformation of normal human erythrocytes. Ex vivo erythrocyte porphyrin analysis pointed to a mild depression of erythropoiesis in dystrophic mice that responded to dietary polyunsaturated fatty acids. The discovery of a paraporphyria in patients with hereditary ataxia was extended to in vitro porphyrin studies using 3-acetylpyridine as an experimental model of ataxia. / Systematic pharmacological studies in a newly designed, serum-free liver cell culture model and in ovo, together with chromatographic-fluorometric PG analysis, established that a certain rate or pattern of PG biosynthesis was required for optimal induction of hepatic (delta)-aminolevulinate synthase and porphyrin-heme biosynthesis. Moreover, the dramatic accumulation of protoporphyrin caused by putative, selective inhibitors of thromboxane synthase, and the exacerbation of experimental porphyria by phospholipase inhibitors provided unique evidence for a physiological, regulatory role of PGs and possibly TXA(,2) in heme biosynthesis at the level of mitochondrial ferrochelatase activity. / Original data obtained about PG effects on heme biosynthesis and erythrocyte properties may prove useful in devising effective therapies and monitoring clinical responses in anemias, porphyrias and hematological aspects of neuromuscular disease.

The capability of axons to grow into a peripheral histoincompatible environment, to locate and to functionally innervate their target structures, was examined through documentation of form and function in nonrejected and rejected upper-extremity composite tissue allografts in adult baboons; Papio h. anubis. Immunosuppression with cyclosporine (radioimmunoassay levels in serum ranging from 1000 to 1200 ng/ml, 12 h after intramuscular injection), supplemented with low doses of methylprednisolone (4.4 mg/day) was sufficient to achieve long-term survival of seven neurovascular free flaps and two hand transplants. / Electrophysiological recordings of more than 600 single axons, combined with light microscopy of the target tissues, revealed that muscle and most classes of skin sensory mechanoreceptors within the allografted tissues became reinnervated. Axons which served allografted tissues that had undergone repeated episodes of rejection showed a significant decrease in conduction velocity, and smaller receptive fields of irregular distribution. Processes of allograft rejection, more than any other factor, led to altered axonal response characteristics.

A decade ago Bizzi and collaborators proposed the "linear summation" hypothesis mainly based on the vestibulo-ocular reflex to explain how subjects move their eyes and head simultaneously to acquire a peripheral visual target. We asked human subjects to make rapid horizontal gaze shifts to unpredictable visible or remembered targets situated at offsets ranging from 30$ sp circ$ to 160$ sp circ$. Different experimental paradigms were used to dissociate crucial variables and different perturbations were imposed to our subjects' head movements to test the interaction between the saccadic eye movement and the head displacement. Several assumptions of the "linear summation" hypothesis were questioned by the analysis of results and we proposed a more elaborated gaze control model to explain how subjects could reach a target accurately with their gaze shifts whatever the conditions, even when the target was beyond the subject's oculomotor range. / In parallel with human experiments we developed a technique to record vestibular units in head-free cats during orienting eye-head movements. Preliminary results on several electrophysiologically identified cells show some particular discharge patterns in the head-free condition. They bring some insights to the behavioral results.

In the feline cerebral cortex flurazepam and ethanol potenitated the inhibitory effects of iontophoretically-applied GABA and electrically-evoked inhibition (believed to be mediated by GABA). This effect is specific, since both flurazepam and ethanol antagonized the inhibitory effects of serotonin, dopamine and glycine. Moreover the degree of potentiation of GABA-mediated electrically-evoked cortical inhibition produced by five benzodiazepines (Ro 21-3981, flurazepam, chlordiazepoxide, medazepam and clozapine): (a) correlated with their relative affinities for the benzodiazepine receptor when all five drugs were applied with equal iontophoretic doses; (b) was not significantly different from one drug to another, when the benzodiazepines were applied in doses inversely proportional to their relative affinities for the benzodiazepine receptor. / Iontophoretically-applied flurazepam consistently potentiated the conductance increase produced by iontophoretically-applied GABA in CA1 and CA3 pyramidal neurons in rat hippocampus. Furthermore, both iontophoretically-applied flurazepam and intravenously injected ethanol consistently prolonged the time course of the GABA-mediated IPSP conductance increase evoked in CA1 and CA3 pyramidal hippocampal neurons by ipsilateral entorhinal or fimbrial stimulation. These effects were independent of membrane potential changes. / It is concluded that GABAergic neurotransmission has a special role in the neurophysiology of anxiety. Since anxiety is involved in the etiology of alcoholism, and since chronic ethanol intake decreases GABA levels and alters the density of GABA receptors, the results of this thesis are consistent with the hypothesis that GABAergic mechanisms may be involved in the etiology of alcoholism.

Atrioventricular (AV) nodal conduction and the pathways of propagation through the AV node have remained enigmatic. Histological and immunohistochemical studies have revealed distinct cellular regions within the AV node with varying distribution of specific ion channels. However, to date, there is little information concerning the pattern of localization of the gap junction proteins that mediate intercellular communication. The current immunohistochemical/confocal studies show a non-uniform distribution of three connexin isoforms (Cx 40, Cx 43 and Cx 45) within these cellular regions of the rat AV node with a marked transition from Cx 43 to Cx 40 at the compact node region. In addition, a high resolution mapping system coupled with extracellular recording electrodes was implemented during this study and has allowed for a macroscopic overview of the rabbit AV node. This approach revealed a marked delay in conduction at the area of the compact node. This information in association with the immunohistochemical studies may help to provide further insight into the mechanisms underlying the AV nodal delay.