Effects of cell concentration during cryopreservation on the post-thaw quality of Santa ines ram sperm.

BACKGROUND: Non-surgical artificial insemination techniques for sheep may benefit from larger numbers of sperm in the insemination dose because the ewe cervix is convoluted and often cannot be traversed with an insemination gun resulting in deposition of the sperm at the os cervix. OBJECTIVE: To compare a range of sperm concentrations when cryopreserving semen from Santa Ines rams and determine the effects of this on post-thaw quality. MATERIALS AND METHODS: One ejaculate from each ram (n = 10) was diluted to four sperm concentrations to obtain the following groups: G-400, G-800, G-1200, and G-1600 x 106 sperm/mL. The semen samples were packaged in 0.25 mL straws, cooled to 5 degree C, cryopreserved in liquid nitrogen vapor, thawed in a water bath (40 degree C per 20 s), and were analyzed for computerized kinetics, capacitation and acrosome integrity, and plasma membrane integrity of sperm. RESULTS: The G-400 treatment resulted in samples with the highest linearity and progressive motion (P < 0.05) and had significantly greater plasma membrane integrity, and lower capacitation and acrosome reaction rates compared to G-1600 (P < 0.05). Overall, use of the G-400 treatment resulted in better kinetics, less plasma membrane damage and less early capacitation. However, despite reducing the ejaculate yield and increasing the costs of the semen freezing process, the G-800 and G-1200 treatments make a greater absolute number of sperm with good kinetics, plasma membrane integrity and capacitation status available. CONCLUSION: Ram sperm concentration impacts cryopreservation, and higher concentrations may be advantageous if a single artificial insemination protocol is desirable. doi.org/10.54680/fr22610110812.

Glyceroneogenesis in the hepatopancreas of the crab Neohelice granulata: Diet, starvation and season effects.

We determined the activity of glyceroneogenesis from [2-14C]-pyruvate, the phosphoenolpyruvate carboxykinase activity, [2-14C]-pyruvate oxidation and total lipid levels in the hepatopancreas of the crab Neohelice granulata fed with a carbohydrate-rich (HC) diet or a high-protein (HP) diet and then subjected to 5weeks of starvation, in summer and winter, to determine whether the seasonal adjustments of lipid metabolism to food scarcity are modulated by the composition of the diet previously given to the crabs. The results demonstrated that glyceroneogenesis is an active pathway in N. granulata hepatopancreas, and is regulated by seasonal variations, diet composition and starvation. This study showed that in summer the increase in the hepatopancreas glyceroneogenesis activity is among the strategies used by N. granulata fed an HP diet, to maintain the triglyceride/fatty acid cycle during starvation, a normal condition in the biological cycle of this crab. However, the administration of an HC diet reduced the glyceroneogenesis capacity in response to starvation in summer. In winter, the decrease in the glyceroneogenesis capacity in both fed (HP and HC diets) and starved crabs seems to be a strategy to reduce energy consumption and/or requirement. In contrast to the summer results, the incorporation of [2-14C]-pyruvate into 14CO2 was markedly higher in both diet (HC and HP) groups and in starved crabs during the winter. Four decades after the first study describing the glyceroneogenesis pathway in rat white adipose tissue, this pathway is evidenced for the first time in a crustacean.

Correlation and path analysis of biomass sorghum production.

Sorghum biomass is an interesting raw material for bioenergy production due to its versatility, potential of being a renewable energy source, and low-cost of production. The objective of this study was to evaluate the genetic variability of biomass sorghum genotypes and to estimate genotypic, phenotypic, and environmental correlations, and direct and indirect effects of seven agronomic traits through path analysis. Thirty-four biomass sorghum genotypes and two forage sorghum genotypes were cultivated in a randomized block design with three replicates. The following morpho-agronomic traits were evaluated: flowering date, stem diameter, number of stems, plant height, number of leaves, green mass production, and dry matter production. There were significant differences at the 1% level for all traits. The highest genotypic correlation was found between the traits green mass production and dry matter production. The path analysis demonstrated that green mass production and number of leaves can assist in the selection of dry matter production.

Photocytotoxicity of a cyanine dye with two chromophores toward melanoma and normal cells.

BACKGROUND: Due to high optical absorption, triplet quantum yield and affinity to biological structures bichromophoric cyanine dyes (BCDs) can be considered promising sensitizers for application in photodynamic therapy (PDT). In this work, we report on the study of the BCD photocytotoxicity toward melanoma and normal cells in comparison with that of commercial photosensitizer Photogem®. METHODS: The cytotoxic and phototoxic effects were measured by standard tests of cell viability. The drug uptake was obtained by the flow cytometry and optical absorption techniques. The BCD intracellular distribution was obtained by the fluorescence image microscopy using specific organelle markers. RESULTS: Both drugs demonstrated increased cytotoxicity under irradiation, while in darkness their cytotoxic effect at concentrations lower than 20 µM after 24 h of incubation did not exceed 20%. For 5 h of incubation, BCD photocytotoxicity in relation to melanoma cells reached 100% already at concentrations below 5 µM, while for normal cells the effect did not exceed 70% even for the 20 µM concentration. It is shown that BCD penetrates into the cells and is located predominantly in perinuclear cytoplasmic structures. CONCLUSIONS: The BCD photosensitizing characteristics appear more adequate for application in PDT than that of the actually applied commercial photosensitizer Photogem®. Higher light absorption by BCD in the near IR region and its preferential localization in mitochondria can explain its high photocytotoxicity. GENERAL SIGNIFICANCE: BCD can be considered as a new promising photosensitizer class for cancer PDT.

Profile of nucleotide catabolism and ectonucleotidase expression from the hippocampi of neonatal rats after caffeine exposure.

Nucleotides and nucleosides play an important role in neurodevelopment acting through specific receptors. Ectonucleotidases are the major enzymes involved in controlling the availability of purinergic receptors ligands. ATP is co-released with several neurotransmitters and is the most important source of extracellular adenosine by catabolism exerted by ectonucleotidases. The main ectonucleotidases are named NTPDases (1-8) and 5'-nucleotidase. Adenosine is a powerful modulator of neurotransmitter release. Caffeine blocks adenosine receptor activity as well as adenosine-mediated neuromodulation. Considering the susceptibility of the immature brain to caffeine and the need for correct purinergic signaling during fetal development, we have analyzed the effects of caffeine exposure during gestational and lactational periods on nucleotide degradation and ectonucleotidase expression from the hippocampi of 7-, 14- and 21-days-old rats. Nucleotides hydrolysis was assessed by colorimetric determination of inorganic phosphate released. Ectonucleotidases expression was performed by RT-PCR. ATP and ADP hydrolysis displayed parallel age-dependent decreases in both control and caffeine-treated groups. AMP hydrolysis increased with caffeine treatment in 7-days-old rats (75%); although there was no significant difference in AMP hydrolysis between control (non caffeine-treated) rats and 14- or 21-days caffeine-treated rats. ADP hydrolysis was not affected by caffeine treatment. Caffeine treatment in 7- and 14-days-old rats decreased ATP hydrolysis when compared to the control group (19% and 60% decrease, respectively), but 21-days-treated rats showed an increase in ATP hydrolysis (39%). Expression levels of NTPDase 1 and 5 decreased in hippocampi of caffeine-treated rats. The expression of 5'-nucleotidase was not affected after caffeine exposure. The changes observed in nucleotide hydrolysis and ectonucleotidases expression could promote subtle effects on normal neural development considering the neuromodulatory role of adenosine.

Zebrafish neurotransmitter systems as potential pharmacological and toxicological targets.

Recent advances in neurobiology have emphasized the study of brain structure and function and its association with numerous pathological and toxicological events. Neurotransmitters are substances that relay, amplify, and modulate electrical signals between neurons and other cells. Neurotransmitter signaling mediates rapid intercellular communication by interacting with cell surface receptors, activating second messenger systems and regulating the activity of ion channels. Changes in the functional balance of neurotransmitters have been implicated in the failure of central nervous system function. In addition, abnormalities in neurotransmitter production or functioning can be induced by several toxicological compounds, many of which are found in the environment. The zebrafish has been increasingly used as an animal model for biomedical research, primarily due to its genetic tractability and ease of maintenance. These features make this species a versatile tool for pre-clinical drug discovery and toxicological investigations. Here, we present a review regarding the role of different excitatory and inhibitory neurotransmitter systems in zebrafish, such as dopaminergic, serotoninergic, cholinergic, purinergic, histaminergic, nitrergic, glutamatergic, glycinergic, and GABAergic systems, and emphasizing their features as pharmacological and toxicological targets. The increase in the global knowledge of neurotransmitter systems in zebrafish and the elucidation of their pharmacological and toxicological aspects may lead to new strategies and appropriate research priorities to offer insights for biomedical and environmental research.

New nitric oxide donors based on ruthenium complexes.

Nitric oxide (NO) donors produce NO-related activity when applied to biological systems. Among its diverse functions, NO has been implicated in vascular smooth muscle relaxation. Despite the great importance of NO in biological systems, its pharmacological and physiological studies have been limited due to its high reactivity and short half-life. In this review we will focus on our recent investigations of nitrosyl ruthenium complexes as NO-delivery agents and their effects on vascular smooth muscle cell relaxation. The high affinity of ruthenium for NO is a marked feature of its chemistry. The main signaling pathway responsible for the vascular relaxation induced by NO involves the activation of soluble guanylyl-cyclase, with subsequent accumulation of cGMP and activation of cGMP-dependent protein kinase. This in turn can activate several proteins such as K+ channels as well as induce vasodilatation by a decrease in cytosolic Ca2+. Oxidative stress and associated oxidative damage are mediators of vascular damage in several cardiovascular diseases, including hypertension. The increased production of the superoxide anion (O2-) by the vascular wall has been observed in different animal models of hypertension. Vascular relaxation to the endogenous NO-related response or to NO released from NO deliverers is impaired in vessels from renal hypertensive (2K-1C) rats. A growing amount of evidence supports the possibility that increased NO inactivation by excess O2- may account for the decreased NO bioavailability and vascular dysfunction in hypertension.

[Acute paraplegia secondary to painless aortic dissection]. / Paraplejía aguda secundaria a la disección indolora de la aorta.

INTRODUCTION: Acute aortic dissection (AAD) is a serious and uncommon event. The clinical presentation generally includes thoracic or back pain. Painless aortic dissection is an extremely rare occurrence. Acute paraplegia is one of the neurological complications secondary to AAD. Although painful paraplegia is seen in 2% to 3% of AAD cases, painless paraplegia is a very rare event. CASE REPORT: A 51-year-old man with a long term history of hypertension, presented with acute paralysis of the lower extremities, with no chest or back pain. CONCLUSION: In presence of acute neurologic events, must always be investigated a vascular cause and, in these group of disease, the aortic dissection may be researched, although the pain wasn't present on occasion of the diagnosis.

Vitamin C improves the effect of a new nitric oxide donor on the vascular smooth muscle from renal hypertensive rats.

Impaired relaxation induced by the new nitric oxide (NO) donor [Ru(NH.NHq)(terpy)NO(+)](3+) (TERPY) has been observed in the aortic rings from renal hypertensive rats (2K-1C). An increased production of reactive oxygen species (ROS) in the aortas from 2K-1C rats are capable of reducing NO bioavailability. Therefore, this study aimed at investigating the effects of an antioxidant (vitamin C) on the relaxant effect of NO released from TERPY on the 2K-1C rat aorta. As for vascular reactivity, the potency of TERPY is greater in the control rats (2K) than in 2K-1C whereas the maximum relaxation (ME) is not significantly different between the 2K and 2K-1C rat aortas. The relaxation of TERPY is potentiated only in the 2K-1C aortic ring treated with vitamin C. TERPY has a lower effect in decreasing cytosolic Ca(2+) concentration ([Ca(2+)]c) in vascular smooth muscle cells (VSMCs) from 2K-1C rats. This effect is also potentiated in 2K-1C aortic cells treated with vitamin C, but it is not altered in 2K cells. The basal cytosolic NO concentration ([NO]c) is lower in 2K-1C than in 2K cells, and the bioavailability of the NO released from TERPY is larger in 2K than in 2K-1C VSMCs. The superoxide radical concentration ([O(2)(*-)]) is higher in the 2K-1C aorta, and vitamin C reduces the [O(2)(*-)] in the 2K-1C aorta. Taken together, these results show that in the aortas of renal hypertensive 2K-1C rats, released NO from the new NO donor is not available to produce a similar effect in 2K aorta due to increased [O(2)(*-)].

Caveolae dysfunction contributes to impaired relaxation induced by nitric oxide donor in aorta from renal hypertensive rats.

Relaxation induced by nitric oxide (NO) donors is impaired in renal hypertensive two kidney-one clip (2K-1C) rat aortas. It has been proposed that caveolae are important in signal transduction and Ca2+ homeostasis. Therefore, in the present study we investigate the integrity of caveolae in vascular smooth muscle cells (VSMCs), as well as their influence on the effects produced by NO released from both the new NO donor [Ru(NH.NHq) (terpy)NO+]3+ (TERPY) and sodium nitroprusside (SNP) on 2K-1C rat aorta. The potency of both TERPY and SNP was lower in the 2K-1C aorta that in the normotensive aorta [two kidney (2K)], whereas the maximal relaxant effect (ME) was similar in both 2K-1C and 2K aortas. In the 2K aorta, methyl-beta-cyclodextrin (CD) reduced both the potency of TERPY and SNP, and their ME compared with the control, but it had no effect on the potency and ME of these NO donors in 2K-1C aortas. The decrease in cytosolic Ca2+ concentration ([Ca2+]c) induced by TERPY was larger in 2K than in 2K-1C cells, and this effect was inhibited by CD in 2K cells only. Aortic VSMCs from 2K rats presented a larger number of caveolae than those from 2K-1C rats. Treatment with CD reduced the number of caveolae in both 2K and 2K-1C aortic VSMCs. Our results support the idea that caveolae play a critical role in the relaxant effect and in the decrease in [Ca2+]c induced by NO, and they could be responsible for impaired aorta relaxation by NO in renal hypertensive rats.

Hepatopancreas gluconeogenesis during hyposmotic stress in crabs Chasmagnathus granulata maintained on high-protein or carbohydrate-rich diets.

The incorporation of [14C]-alanine or [14C]-lactate into glucose was measured in hepatopancreas fractions from Chasmagnathus granulata crabs adapted to a high protein or a carbohydrate-rich diet and submitted or not (control group) to hyposmotic stress. Gluconeogenic capacity and phosphoenolpyruvate carboxykinase (PEPCK) activity increased during acclimation to a dilute medium in C. granulata hepatopancreas. In intact animals, high hemolymph urea levels occurred for the high-protein regimen and for crabs fed both diets and submitted to hyposmotic stress. It could be that the amino acids released during hyposmotic stress are deaminated in the hepatopancreas, and that the carbon chains are used as substrate for gluconeogenesis. Hepatopancreas gluconeogenesis seems to be one of the pathways implicated in the metabolic adjustment of the amino acid pool during hyposmotic stress in C. granulata.

[Functional properties and protein concentrate of Pigeon pea (Cajanus cajan (I.) Millsp) flour]. / Propriedades funcionais da farinha e concentrado protéico de feijão guandu (Cajanus cajan (I.) Millsp).

The objective of this investigation was to study the functional properties of Pigeon pea (Cajanus cajan (L.) Millsp) flour and protein concentrate. The solubility of both samples were superior than 70% at pH above 6.7 and below 3.5. The water and oil absorption were 1.2 and 1.07 ml/g of sample and 0.87 and 1.73 ml/g of flour and protein concentrate samples, respectively. The minimum concentration of flour and protein concentrate needed for gelation was 20% and 12%, respectively. The emulsifying capacity of flour and concentrate was 129.35 g and 191.66 g oil/g of protein and the emulsion stability 87.50 and 97.97%, respectively, after 780 minutes. The foam capacity and stability of flour foam were 36.0% and 18.61, while of the concentrate were 44.70% and 78.97% after 90 minutes. These properties indicate that the flour as well as the concentrate could have application in various food systems.

Pharmacokinetics of radioiodinated growth hormones in the turtle Chrysemys dorbigni.

Growth hormone binding proteins (GHBP) have been identified in the blood of many species. The aim of the present work is to study the physiological role of the GHBP in the turtle serum which we recently described. Binding studies were carried out using in vivo pharmacokinetic and chromatographic techniques as well as in vitro methods. When (125)I-GH was injected in physiological concentration into Chrysemys dorbigni turtles, the first step of pharmacokinetics was the binding of a significant fraction of the labeled GH by the GHBPs present in serum. The decay curve followed a three compartments model and gave the equation: Ae(-alphat) + Be(-betat) + Ce(-gammat). The fast compartment with t(1/2) of 14.4 min or 25.2 min, for hGH and bGH represents 30.3% and 18.9% of total radioactivity, respectively, at hypothetical time zero (not experi mental). Chromatographic studies reveal that this rapid compartment represents free GH. The second and third compartments represent complex forms between GH and GHBPs present in the turtle serum, and represent 70% and 80% of total radioactivity for hGH and bGH, respectively. In vitro chromatographic studies showed direct evidence of the presence of GHBPs in the turtle serum. The presence of these GHBPs changed the pharmacokinetics of labeled GH in plasma and the subsequent liver uptake of GH. The labeled hGH or bGH binds to turtle serum in similar proportion, but maximal liver uptake of these hormones are completely different (L/B ratio of 9.2 +/- 0.6 (n = 5) for ( 125)I-hGH and 4.8 +/- 0.3 (n = 7) for (125)I-bGH). The reasons for these differences could be that human GH binds to lactogenic and somatotropic receptors and bovine GH binds only to somatotropic receptors.

Insulin binding sites in gills of the estuarine crab Chasmagnathus granulata.

Bovine 125I-insulin was injected into the estuarine crab Chasmagnathus granulata in order to study its distribution and specific uptake by tissues. The highest radioactivity uptake occurred in both anterior and posterior gills, which reached maximum values at 30-60 min following labeled insulin administration. Heart and hepatopancreas concentrated a very low amount of radioactivity (only 9 and 3%, respectively, of that shown by gills). A significant reduction of the uptake was observed in the gills when an excess of unlabeled insulin was injected together with the labeled hormone. In vitro studies also showed specific uptake of 125I-insulin by the gills incubated at 25 degrees C, which reached a plateau after 120-min incubation, suggesting a saturable process. The inhibition of 125I-insulin uptake was dose dependent on unlabeled insulin. Glucagon did not compete with radioactivity uptake by gills in vivo and in vitro. Further characterization of insulin-binding sites was performed in gill membrane. The amount of unlabeled insulin that prevented 50% of the 125I-insulin uptake was 7.78 micrograms/ml, and the Scatchard plot analysis established the presence of binding site with Kd of 3.11 microM and Bmax of 0.14 microM (r = 0.99). Ovine prolactin was not able to prevent. 125I-insulin binding to gill membrane. These findings seem to indicate the presence of specific binding sites for insulin or insulin-like substance in crab gills, which deserves further studies.