Non-Selective Calcium Channel Blocker Bepridil Decreases Secondary Pathology in Mice after Photothrombotic Cortical Lesion

Experimental studies have identified a complex link between neurodegeneration, β-amyloid (Aβ) and calcium homeostasis. Here we asked whether early phase β-amyloid pathology in transgenic hAPP_SL mice exaggerates the ischemic lesion and remote secondary pathology in the thalamus, and whether a non-selective calcium channel blocker reduces these pathologies. Transgenic hAPP_SL (n = 33) and non-transgenic (n = 30) male mice (4–5 months) were subjected to unilateral cortical photothrombosis and treated with the non-selective calcium channel blocker bepridil (50 mg/kg, p.o., once a day) or vehicle for 28 days, starting administration 2 days after the operation. Animals were then perfused for histological analysis of infarct size, Aβ and calcium accumulation in the thalamus. Cortical photothrombosis resulted in a small infarct, which was associated with atypical Aβ and calcium accumulation in the ipsilateral thalamus. Transgenic mice had significantly smaller infarct volumes than non-transgenic littermates (P<0.05) and ischemia-induced rodent Aβ accumulation in the thalamus was lower in transgenic mice compared to non-transgenic mice (P<0.01). Bepridil decreased calcium load in the thalamus (P<0.01). The present data suggest less pronounced primary and secondary pathology in hAPP_SL transgenic mice after ischemic cortical injury. Bepridil particularly decreased calcium pathology in the thalamus following ischemia.     

Hormonal regulation of myosin heavy chain and alpha-actin gene expression in cultured fetal rat heart myocytes

Thyroid hormone regulates the expression of ventricular myosin isoenzymes by causing an accumulation of alpha-myosin heavy chain (MHC) mRNA and inhibiting expression of beta-MHC mRNA. However, the mechanism of thyroid hormone action has been difficult to examine in vivo because of its diverse actions. Accordingly, hormonal control of expression of six MHC isoform mRNAs and cardiac and skeletal alpha-actin mRNAs was studied in primary cultures of fetal rat heart myocytes grown in defined medium. The results indicate that in the absence of thyroid hormone, cultured heart cells express predominantly beta-MHC and cardiac alpha-actin mRNAs. Addition of 3,5,3'-triiodo-L-thyronine (T3) caused a rapid induction of alpha-MHC mRNA and decreased beta-MHC mRNA levels without affecting the skeletal muscle MHC mRNAs. There was an almost parallel change in the myosin isoenzymes. Cardiac alpha-actin mRNA levels were transiently increased by T3 treatment, but skeletal alpha-actin was unaffected. Elimination of insulin and epithelial growth factor from the medium did not alter the effects of T3 on cardiac MHC mRNA expression. Addition of various adrenergic agents to the medium had no appreciable effect on cardiac MHC mRNA expression despite the presence of functionally coupled alpha- and beta-adrenergic receptors. Addition of steroid hormones, muscarinic agents, and glucagon to the medium also had no effect. Thus, under defined conditions, T3 is able to regulate MHC gene expression at a pretranslational level without the need for other exogenous factors.     

Changes in Leydig cell activity during the annual testicular cycle of the bat Myotis lucifugus lucifugus: histology and lipid histochemistry

Changes in Leydig cell histology and testicular sudanophilic lipids were examined in relation to spermatogenic activity in the bat Myotis lucifugus lucifugus (Chiroptera: Vespertilionidae) throughout the annual cycle in the northeastern United States. These changes were correlated with annual variations in plasma testosterone concentrations which have recently been described for this species. Gametogenic activity occurred during the months of May-August when bats were metabolically most active. During hibernation (October-April), when sperm are stored in the epididymides, and accessory glands are hypertrophic, the seminiferous tubules were at rest, and the germinal epithelium was reduced to reserve spermatogonia and Sertoli cells. Based on their structure and cyclic pattern of sudanophilic lipids, Leydig cells exhibited a pattern of activity that closely paralleled that of the seminiferous epithelium. On renewal of spermatogenesis in spring, Leydig cells became hypertrophied and accumulated lipid inclusions. These inclusions, seen as vacuoles in plastic sections and sudanophilic droplets in frozen sections, reached maximal accumulations in late June. In late July and during August, when peak testosterone levels occur in blood, lipid droplets were dramatically depleted, and Leydig cells were weakly sudanophilic. In September, when testosterone titers return to low baseline levels, Leydig cells had regressed but exhibited a marked increase in sudanophilic inclusions which appeared to be mostly lipofuscins. During the ensuing mating and hibernation periods, Leydig cells were involuted and filled with lipofuscins. During the periarousal period, however, Leydig cells became weakly Sudan-positive while many large, intensely sudanophilic cells were scattered throughout the interstitium. In electron micrographs these cells were identified as macrophages. They appear to play an important role in the annual testicular cycle by phagocytizing the residues of Leydig cell involution in preparation for a new steroidogenic cycle. Seasonal changes in lipid inclusions were also observed in the seminiferous tubules. In addition, the relationship of the Leydig cell cycle to androgen action and the accessory organs in this bat is discussed.     

Molecular characterization of the diurnal/circadian expression of the chlorophyll a/b-binding proteins in leaves of tomato and other dicotyledonous and monocotyledonous plant species

Diurnal oscillations of steady-state mRNA levels encoding the chlorophyll a/b-binding proteins were monitored inLycopersicon esculentum, Glycine max, Phaseolus vulgaris, P. aureus, P. coccineus, Pisum sativum, Sinapis alba, Hordeum vulgare, Triticum aestivum andZea mays. In these plant speciescab mRNA accumulation increases and decreases periodically indicating i) that the expression of the genes for chlorophyll a/b-binding proteins (cab genes) is controlled by a circadian rhythm, and ii) that the rhythm is widely distributed among monocotyledonous and dicotyledonous plant species. A detailed characterization of the pattern ofcab mRNA expression in tomato leaves shows that the amplitude of the oscillation is dependent on i) the developmental stage of the leaves, ii) the circadian phase and duration of light and iii) the circadian phase and duration of darkness. In addition to the chlorophyll a/b-binding proteins, genes coding for other cellular functions were examined for cyclic variations of their mRNA levels. The analysis includes genes involved in i) carbon metabolism (e.g. phosphoenolpyruvate carboxylase, pyruvate orthophosphate dikinase, alpha amylase, fructose-1,6-bisphosphate aldolase, triosephosphate isomerase), ii) photosynthesis (large and small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase, Q_B-binding protein, reaction-center protein of photosystem I) and iii) other physiological or morphological reactions (e.g. ubiquitin, actin). However, no periodic fluctuation pattern was detected for the mRNA levels of these genes in tomato and maize leaves.     

An improved method for light- and electron microscopial studies of nematode/fungal interactions

A method is presented that enables studies to be made of single nematode-fungal interactions under conditions where fungal growth at the expense of external nutrients is prevented. The nematophagous fungus Arthrobotrys ologospora was used as a model organism in these studies. The method is based on removal of the traps from the vegetative mycelium, immediately after a nematode was captured and transfer of the trap with the captured nematode into a droplet of sterile distilled water placed in a moisture chamber. In the absence of external nutrients, such isolated traps of A. oligospora were fully effective in penetrating and subsequently digesting the captured nematode. Solely vegetative mycelium was formed at the expense of the digested nematode; this developed from the trap that originally had captured the nematode. One advantage of the present method is that studies on various stages of the nematode-fungal interaction can now be performed under conditions that exclude major influences of external nutrients which otherwise could be communicated to the trap cells by way of the vegetative mycelium.     

Hyphal fusion during initial stages of trap formation in Arthrobotrys oligospora

Hyphal fusion during initial stages of trap formation by Arthrobotrys oligospora was studied by video-enhanced contrast and electron microscopy. Trap initials grew perpendicularly to the parent hypha, then curved around and anastomosed with a peg that developed on the hypha. Trap initials usually developed 40–140 m apart while the anastomosis occurred 20–25 m from the initial. Vigorous cytoplasmic movements in trap initials and developed traps corresponded to intense staining with fluorescein diacetate (FDA) of these cells. In addition, bundles of microfilaments were seen in developing loops of traps. On fusion organelle migration took place from the tip cell of the trap into the peg. Later on a septum was formed at the site of fusion.