Increased α1 receptor density in brown adipose tissue of cafeteria-fed rats

A possible general corollary between ₁-receptor density in brown adipose tissue and the degree of activation of the tissue was investigated. For this purpose, the effect of cafeteria feeding on ₁-adrenergic receptors in brown adipose tissue of seven-week-old female rats was studied by the use of the ₁-antagonist (³H)prazosin. In cafeteria-fed rats, the K_D of the ₁-receptor for (³H)prazosin was unchanged (about 0.35 nM), but the receptor density was doubled (up to 40 fmol per mg of membrane protein). This was also observed when the results were expressed per unit of a plasma-membrane marker (5-nucleotidase). It was concluded that an increased ₁-receptor density is seen not only in cold-acclimated rats, but also in other conditions where brown fat is activated, and a possible general physiological significance of ₁-adrenergic pathways in brown adipose tissue is discussed.     

Pharmacological characterization of alpha1- and beta-adrenergic synergism of 5'DII activity in rat brown adipocytes

The role of adrenoceptor subtypes was studied in rat brown adipose tissue (BAT). The type II 5'-deiodinase (5'DII) was activated in response to simultaneous stimulation by beta3- and alpha1-adrenergic agonists, BRL 37344 or CGP 12177, and cirazoline, in brown adipocytes. Inhibition of the alpha1- and beta-adrenergic phenylephrine-stimulated 5'DII activity was obtained by the alpha1-adrenergic antagonists in the order of prazosin >/= wb 4101 > 5-methylurapidil. In comparison, the binding of [3H]prazosin to rat BAT plasma membranes was inhibited by alpha1-adrenergic antagonists in the order of prazosin > WB 4101 = benoxathian > 5-methylurapidil. Although the order of the alpha1-adrenergic competition seemed to be rather typical for the alpha1B-adrenergic receptors, a molecular analysis on adrenoceptor mRNAs should be made to confirm the exact alpha1-adrenergic subtypes at the level of brown adipocytes, since the possibility of a mixture of different receptor subtypes in brown fat cells and/or tissue may interact with the pharmacological characterization. Thus, specific alpha1- and beta-adrenoceptor subtypes participate in the regulation of 5'DII activity in the rat brown adipocytes, and therefore, an impaired alpha1- and beta-adrenergic co-work may be involved in a defective BAT function, e.g., in obese Zucker rats, too. An interesting possibility is that the decreased number of alpha1-adrenoceptors in the BAT of obese Zucker rats is due to the decrease in the alpha1B-adrenoceptor subtype which would further be involved especially in the regulation of BAT 5'DII activity.     

Catechol-O-Methyltransferase (COMT) Protein Expression and Activity after Dopaminergic and Noradrenergic Lesions of the Rat Brain

The occurrence of catechol-O-methyltransferase (COMT) in presynaptic neurons remains controversial. This study utilized dopaminergic and noradrenergic toxins to assess the presence of COMT in the presynaptic neurons originating from the substantia nigra, ventral tegmental area or locus coeruleus. Destruction of dopaminergic and noradrenergic neurons was assessed by measuring the dopamine and noradrenaline content in the projection areas of these neurons. Additionally, COMT protein expression and activity were examined in several projection areas to determine whether there are any changes in COMT values. Colocalization studies were done to identify COMT-containing postsynaptic neurons. Despite successful lesioning of dopaminergic and noradrenergic neurons, no changes in COMT protein expression or activity could be noted. These results strongly suggest that COMT is not present in presynaptic dopaminergic and noradrenergic neurons. There was a high colocalization of COMT with the GABAergic marker of short neurons both in the striatum and cortex but only a weak, if any, with the cholinergic marker in the cortex.     

Small Tails Tell Tall Tales – Intra-Individual Variation in the Stable Isotope Values of Fish Fin

Background

Fish fin is a widely used, non-lethal sample material in studies using stable isotopes to assess the ecology of fishes. However, fish fin is composed of two distinct tissues (ray and membrane) which may have different stable isotope values and are not homogeneously distributed within a fin. As such, estimates of the stable isotope values of a fish may vary according to the section of fin sampled.

Methods

To assess the magnitude of this variation, we analysed carbon (δ ¹³C), nitrogen (δ ¹⁵N), hydrogen (δ ²H) and oxygen (δ ¹⁸O) stable isotopes of caudal fin from juvenile, riverine stages of Atlantic salmon (Salmo salar) and brown trout (Salmo trutta). Individual fins were sub-sectioned into tip, mid and base, of which a further subset were divided into ray and membrane.

Findings

Isotope variation between fin sections, evident in all four elements, was primarily related to differences between ray and membrane. Base sections were¹³C depleted relative to tip (~ 1 ‰) with equivalent variation evident between ray and membrane. A similar trend was evident in δ ²H, though the degree of variation was far greater (~ 10 ‰). Base and ray sections were ¹⁸O enriched (~ 2 ‰) relative to tip and membrane, respectively. Ray and membrane sections displayed longitudinal variation in ¹⁵N mirroring that of composite fin (~ 1 ‰), indicating that variation in¹⁵N values was likely related to ontogenetic variation.

Conclusions

To account for the effects of intra-fin variability in stable isotope analyses we suggest that researchers sampling fish fin, in increasing priority, 1) also analyse muscle (or liver) tissue from a subsample of fish to calibrate their data, or 2) standardize sampling by selecting tissue only from the extreme tip of a fin, or 3) homogenize fins prior to analysis.     

Basin‐scale phenology and effects of climate variability on global timing of initial seaward migration of Atlantic salmon (Salmo salar)

Migrations between different habitats are key events in the lives of many organisms. Such movements involve annually recurring travel over long distances usually triggered by seasonal changes in the environment. Often, the migration is associated with travel to or from reproduction areas to regions of growth. Young anadromous Atlantic salmon (Salmo salar) emigrate from freshwater nursery areas during spring and early summer to feed and grow in the North Atlantic Ocean. The transition from the freshwater (‘parr’) stage to the migratory stage where they descend streams and enter salt water (‘smolt’) is characterized by morphological, physiological and behavioural changes where the timing of this parr‐smolt transition is cued by photoperiod and water temperature. Environmental conditions in the freshwater habitat control the downstream migration and contribute to within‐ and among‐river variation in migratory timing. Moreover, the timing of the freshwater emigration has likely evolved to meet environmental conditions in the ocean as these affect growth and survival of the post‐smolts. Using generalized additive mixed‐effects modelling, we analysed spatio‐temporal variations in the dates of downstream smolt migration in 67 rivers throughout the North Atlantic during the last five decades and found that migrations were earlier in populations in the east than the west. After accounting for this spatial effect, the initiation of the downstream migration among rivers was positively associated with freshwater temperatures, up to about 10 °C and levelling off at higher values, and with sea‐surface temperatures. Earlier migration occurred when river discharge levels were low but increasing. On average, the initiation of the smolt seaward migration has occurred 2.5 days earlier per decade throughout the basin of the North Atlantic. This shift in phenology matches changes in air, river, and ocean temperatures, suggesting that Atlantic salmon emigration is responding to the current global climate changes.     

Production of functional recombinant tyrosine hydroxylase by the BPV-1 expression plasmids in the cell cultures

Bovine papilloma virus type-1 (BPV-1)-based expression plasmids TkBPVTH and CGalBPVTH encoding the rat tyrosine hydroxylase (TH) enzyme have been designed for the development of gene therapy for experimental Parkinson's disease. The aim of the present work was to examine the transfection of BPVTH plasmids to express a dopaminergic transgene in the monkey CV1-P fibroblast, rat C6 glioma and human NHA astrocyte cell cultures. The biological function of the transgene was estimated by analyzing the production of recombinant TH mRNA and protein, and the synthesis of L-dopa and dopamine. The highest transfection efficiency was obtained using TkBPVTH plasmids (5 microg). Furthermore, the expression of TkBPVTH plasmids was associated with significant synthesis of TH enzyme and L-dopa in the C6 and NHA cell cultures.     

The use of low-molecular-weight PEIs as gene carriers in the monkey fibroblastoma and rabbit smooth muscle cell cultures

Background

Polyethylenimines (PEIs) and cationic polymers have been used successfully in gene delivery. In earlier reports, only large PEIs (MW>10 000) have shown significant transfection efficiency. In the present study, the roles of small PEIs (MW 700 and 2000) were studied as additional compounds to see if they can improve gene delivery with cationic liposomes.

Methods

The TKBPVlacZ expression plasmid was transfected in the CV1‐P (monkey fibroblastoma) and SMC (rabbit smooth muscle) cell lines using various combinations of PEIs (MW 700, 2000, and 25 000) and Dosper liposomes. The transfection efficiency was determined with the fluorometric ONPG (o‐nitrophenol‐β‐D ‐galactopyranoside) assay and histochemical X‐gal staining. The toxicity of the transfection reagents was estimated by the MTT [3‐(4,5‐dimethylthiazolyl‐2)‐2,5‐diphenyl tetrazolium bromide] assay.

Results

Transfection of TKBPVlacZ plasmid by the small PEIs (MW 700 and 2000) combined with Dosper liposomes was associated with high expression of the lacZ reporter gene in the CV1‐P and SMC cell lines. The transfection efficiencies of the low‐molecular‐weight PEI/liposome combinations were several fold higher than those of PEIs or liposomes alone. PEI/liposome combinations had no toxicity on the cell lines tested.

Conclusions

The low‐molecular‐weight PEIs could be used successfully for gene delivery when combined with the cationic liposomes, resulting in a synergistic increase of the transfection efficiency in both cell lines studied. Copyright © 2002 John Wiley & Sons, Ltd.

     

Spawning migration behaviour of sea trout (Salmo trutta L.) in a boreal river system: effects of flow conditions and obstacles on migratory activity

In this study, radio telemetry was used to examine the upstream spawning migration behaviour of anadromous brown trout (sea trout), Salmo trutta L., in a boreal river system, the River Isojoki, western Finland. The aim was to study the movement activity and migration characteristics of trout during the upstream spawning migration, as well as to locate the important spawning habitats and study the spawning characteristics. Furthermore, the authors analysed how flow conditions and a hydropower dam, with adjacent fishways, affected the upstream spawning migration. Tagged trout spawned in both the main stem and four tributaries, with spawning taking place from early October to November. The movement activity of radio-tagged trout was influenced by a hydropower dam (Perus dam), with spring migrators spending prolonged periods at the dam area, postponing the migration upstream. Flow conditions affected the total time spent at the dam area, as well as the movement activity in the free-flowing sections above the dam, with increasing flow stimulating activity. In addition, time of river ascent and location of spawning area had a significant effect on the movement activity of tagged trout. These results are further evidence that synergistic effects of flow and migratory obstacles can negatively influence migrations of anadromous fish, regardless of constructed fishways. The management of flow regimes and the efficiency of fishways are vital, as climate change will likely influence the flow and increase the water temperature of boreal river systems, further aggravating issues caused by obstacles.     

Efficient Transfection of Novel Bovine Papillomavirus 1 Expression Plasmids

A series of novel bovine papillomavirus type 1 (BPV-1)-based expression plasmids was constructed and characterized in vitro as a starting point for the development of an in vivo gene therapeutic method. The order of transfection efficiency for different pBPVlacZ plasmids was pCGalBPV > pTKBPV > pSRalphaBPV in CV1-P cells. In the absence of selection pressure, the expression of pCGalBPVlacZ and pTKBPVlacZ was associated with long-term maintenance. In a comparison of pBPVlacZ with pSVlacZ, expression was maintained up to 12-17 and 8-12 days, respectively. The transfection of pBPVlacZ plasmids was efficient in secondary and primary, dividing and nondividing, neural and nonneural, and human cells and, furthermore, independent of the cell cycle as seen in growing as well as resting cells. All these characteristics are likely to be relevant for in vivo conditions, under which the percentage of proliferating cells could be quite low. In conclusion, the pBPV plasmids were efficiently delivered and expressed in different host cells, and therefore their performance in gene therapy is worth testing.