γ-Amino Butyric Acid Control of Arginine Vasopressin Release from the Ewe Hypothalamus In Vitro: Sensitivity to Oestradiol

The present study aims to ascertain the influence of gamma-amino butyric acid (GABA)(A or B) receptors on arginine vasopressin (AVP) release in vitro and determine whether E(2) modulates GABA-AVP interaction. Within 10 min of ewe killing, saggital midline hypothalamic slices (from the anterior preoptic area to the mediobasal hypothalamus along with the median eminence, 2-mm thick, two per ewe) were dissected, placed in oxygenated minimum essential media (MEM)-alpha at 4 degrees C and within 2 h were singly perifused at 37 degrees C with oxygenated MEM-alpha (pH 7.4; flow rate 0.15 ml/min), either with or without E(2) (24 pg/ml). After 4-h equilibration, 10-min fractions were collected for 4 h interposed with a 10-min exposure at 60 min to a specific GABA(A or B) receptor agonist or antagonist at various doses (0.1-10 mm). GABA(A) (muscimol; no E(2), n = 7 perifusion chambers, with E(2), n = 11) or GABA(B) (baclofen; no E(2), n = 8, with E(2), n = 15) agonists (10 mm) did not influence AVP concentrations. However, AVP release increased (p < 0.05) 20-30 min after exposure to 10 mm GABA(A or B) antagonists (bicuculline, no E(2), n = 7: from 4.6 +/- 0.7 to 33.0 +/- 0.4, with E(2), n = 17: from 11.9 +/- 1.4 to 32.8 +/- 6.0; CGP52432, with E(2), n = 14: from 14.0 +/- 2.6 to 28.8 +/- 3.9 pg/ml). At the end of the collection period, hypothalamic slices responded to KCl (100 mm) with AVP efflux (p < 0.05). GABA(B) but not GABA(A) antagonist-stimulated AVP release was enhanced in the presence of E(2). In summary, AVP release is under the inhibitory influence of GABA input with further potentiation by E(2) through GABA(B) receptors in vitro.     

Linking Land-use, Water Body Type and Water Quality in Southern New Zealand

Land-use and vegetation cover have been linked to the nutrient levels (nitrogen, phosphorus) of surface waters in several countries. However, the links generally relate to streams and rivers, or to specific types of standing water, for example shallow lakes in a geologically defined region. We measured physical variables and nutrient chemistry of 45 water bodies representative of the wide range of lentic wetland environments (swamps, riverine wetlands, estuaries, reservoirs, shallow lakes, deep lakes) in Otago, New Zealand, and related these to catchment variables and land-use in order to assess the potential influence of catchment modification on water quality of these diverse wetlands. Catchment boundaries and land cover were derived from maps using ArcView GIS software. Our predictions that concentrations of nutrients and other components of water quality would correlate positively with the nature and intensity of catchment modification were confirmed in multivariate analyses. Physical and chemical measures were positively related to the extent of modification in the catchment (percentage of the catchment in pasture, planted forest, scrub and urban areas), and negatively related to lack of catchment modification (more of the catchment in bare ground, tussock grassland and indigenous forest). The strong negative correlations between nutrient concentrations, suspended sediment, water colour and the percentage of tussock cover in the catchment imply that increased conversion of the␣native tussock grassland to pastoral farming in␣Otago will increase nutrient concentrations and␣reduce water quality of the diverse lentic ecosystems.     

Immunolocalization of Aquaporins 1 and 9 in the Ram Efferent Ducts and Epididymis

Aquaporins (AQPs) are essential membrane protein channels for the transport of water across membranes. Fluid movement in the epididymis is important for modulation of the luminal environment, in which sperm mature and reside. This study was designed to understand the morphology and localization of AQPs in ram efferent ducts (ED) and epididymis. For this purpose, the epididymis of seven animals were removed for histologic and immunohistochemical analyses. AQP1 immunoreactivity was observed in the apex of the ED, and AQP9 was found adjacent to the nuclei of the epithelial cells of the ED. The epithelial lining of ram epididymis is pseudostratified columnar and presents principal, basal, apical and narrow cells. In the initial segment (IS), a moderate reaction for AQP1 was observed in the apical cytoplasm of epithelial cells. An intense reactivity for AQP1 was noted over the microvilli of principal cells and in spermatozoa in the caput. In the corpus and cauda, AQP1 was noted only over the endothelial cells of vascular channels located in intertubular spaces. A weak‐to‐moderate reaction for AQP9 was observed in the nuclei of epithelial cells in the IS, caput and corpus of the epididymis. In the cauda, an intense reaction to AQP9 was observed in the epithelial border. In the IS, caput and corpus, the reactivity for AQP9 differed from those observed in domestic animals. The cauda showed a pattern similar to that previously described. These results indicate that AQPs 1 and 9 have reversed locations and roles in rams, suggesting activity variations related with fluid and solute absorption throughout the epididymis.     

Kisspeptin,c‐Fos and CRFR Type 2 Co‐expression in the Hypothalamus after Insulin‐Induced Hypoglycaemia

Normal reproductive function is dependent upon availability of glucose and insulin‐induced hypoglycaemia is a metabolic stressor known to disrupt the ovine oestrous cycle. We have recently shown that IIH has the ability to delay the LH surge of intact ewes. In the present study, we examined brain tissue to determine: (i) which hypothalamic regions are activated with respect to IIH and (ii) the effect of IIH on kisspeptin cell activation and CRFR type 2 immunoreactivity, all of which may be involved in disruptive mechanisms. Follicular phases were synchronized with progesterone vaginal pessaries and at 28 h after progesterone withdrawal (PW), animals received saline (n = 6) or insulin (4 IU/kg; n = 5) and were subsequently killed at 31 h after PW (i.e., 3 h after insulin administration). Peripheral hormone concentrations were evaluated, and hypothalamic sections were immunostained for either kisspeptin and c‐Fos (a marker of neuronal activation) or CRFR type 2. Within 3 h of treatment, cortisol concentrations had increased whereas plasma oestradiol concentrations decreased in peripheral plasma (p < 0.05 for both). In the arcuate nucleus (ARC), insulin‐treated ewes had an increased expression of c‐Fos. Furthermore, the percentage of kisspeptin cells co‐expressing c‐Fos increased in the ARC (from 11 to 51%; p < 0.05), but there was no change in the medial pre‐optic area (mPOA; 14 vs 19%). CRFR type 2 expression in the lower part of the ARC and the median eminence was not altered by insulin treatment. Thus, disruption of the LH surge after IIH in the follicular phase is not associated with decreased kisspeptin cell activation or an increase in CRFR type 2 in the ARC but may involve other cell types located in the ARC nucleus which are activated in response to IIH.     

A weight‐of‐evidence approach for defining thermal sensitivity in a federally endangered species

1. Managing for threatened and endangered species under changing environmental conditions is a challenge faced by resource managers worldwide. Lack of basic knowledge of the biology and habitat requirements of these species can contribute to this difficulty but is exacerbated by the limitations of working with rare species (i.e. few individuals) or unrefined and non‐lethal methods for evaluating stress.
2. A weight‐of‐evidence approach was used to evaluate the thermal biology of the federally endangered dwarf wedgemussel (Alasmidonta heterodon), using cumulative results from multiple experimental assessments, co‐occurring species, and their host fish to begin defining the thermal limits and optimal conditions for the species.
3. Results suggest that dwarf wedgemussel and its host fish are thermally sensitive species compared with other Atlantic‐slope mussels, with a lower critical thermal maximum (CTM) and the selection of reduced temperatures during choice experiments.
4. Physiological studies resulted in a lack of statistical significance, primarily owing to low power, which was a function of sample size – an unavoidable problem when studying rare species. Given these limitations, thermal choice and CTM may be more useful end points than physiological processes such as clearance and respiration rates when dealing with sample‐size limitations.
5. These results suggest that management strategies that avoid exposing dwarf wedgemussel and its thermally sensitive host fish to extreme temperatures could be important for species conservation.

     

Pharmacokinetics and clinical efficacy of a cinchophen and prednisolone combination in the dog

The kinetics and efficacy of a cinchophen prednisolone combination preparation (PLT) were assessed in the dog. Cinchophen administered at a dose rate of 12-5 mg/kg intravenously had a volume of distribution (Vd area) of 0–13 litres/kg, a clearance rate (Cl) of 0–15 litres/h and a half-life (t _1/2β of 7–92 hours. Following oral administration the bioavailability was 87-21 per cent. Prednisolone administered at a dose rate of 0–15 mg/kg intravenously had a Vd area of 2–7 litres/kg, a CI of 0–116 litres/h and a t_1/2β of 1–11 hours. PLT given to dogs with osteoarthritis at a dosage range of between 25 and 44 mg/kg per day cinchophen and between 0–125 and 0–220 mg/kg per day prednisolone for a maximum of 14 days significantly improved lameness (P<0–001), weight bearing (P<0–005), joint mobility (P<0–01) and stiffness (P<0–001) scores and was similar in clinical efficacy to phenylbutazone.     

No DNA loss in autotetraploids of Arabidopsis thaliana

To address the issue of genome evolution in autopolyploids and particularly to investigate whether rapid sequence elimination also occurs in autopolyploids as in allopolyploids, amplified fragment length polymorphism (AFLP) fingerprinting was employed to examine a large number of genomic loci in F₁ hybrids between two different autotetraploids of Arabidopsis thaliana accessions, namely Ler and Col. Using this approach, perfect additivity in the F₁ hybrids was found between the newly‐formed autopolyploids when compared with their parental lines. Using flow cytometry, the study was extended in a quantitative manner, in which the nuclear DNA contents in one autotetraploid A. thaliana accession Ler, was determined. The increase in genome size of the autotetraploid line was additive. Taken together, no evidence was found for genome size reduction due to autopolyploidization of A. thaliana. The results indicating that there was no DNA loss in autotetraploid A. thaliana suggest that a different type of genome evolution may occur in autopolyploids during the initial stages of their formation when compared with allopolyploids.     

Rapid flow cytometric analysis of the cell cycle in intact plant tissues

Mechanical chopping of plant tissues in the presence of mithramycin released intact nuclei representative of the cells within the tissues. The amount of nuclear DNA in the homogenates of monocotyledonous and dicotyledonous plants was accurately and rapidly determined by flow microfluorometry, and the distribution of nuclei involved in the cell cycle was charted for tissues selected from different physical locations or developmental stages.