The cannabinoid receptors system in horses: Tissue distribution and cellular identification in skin

BackgroundThe endocannabinoid system (ECS) is composed of cannabinoid receptors type 1 (CBR1) and type 2 (CBR2), cannabinoid‐based ligands (endogenous chemically synthesized phytocannabinoids), and endogenous enzymes controlling their concentrations. Cannabinoid receptors (CBRs) have been identified in invertebrates and in almost all vertebrate species in the central and peripheral nervous system as well as in immune cells, where they control neuroimmune homeostasis. In humans, rodents, dogs, and cats, CBRs expression has been confirmed in the skin, and their expression and tissue distribution become disordered in pathological conditions. Cannabinoid receptors may be a possible therapeutic target in skin diseases.ObjectivesTo characterize the distribution and cellular expression of CBRs in the skin of horses under normal conditions.AnimalsFifteen healthy horses.MethodsUsing full‐thickness skin punch biopsy samples, skin‐derived primary epidermal keratinocytes and dermal‐derived cells, we performed analysis of Cnr1 and Cnr2 genes using real‐time PCR and CBR1 and CBR2 protein expression by confocal microscopy and Western blotting.ResultsNormal equine skin, including equine epidermal keratinocytes and dermal fibroblast‐like cells, all exhibited constant gene and protein expression of CBRs.Conclusions and Clinical ImportanceOur results represent a starting point for developing and translating new veterinary medicine‐based pharmacotherapies using ECS as a possible target.     

Termination of mid-term pregnancy in the dog with oral RU 486

A total of four pregnancies were terminated in three bitches (two beagles and one flatcoated retriever) with a single dose of 20 mg/kg (one case), two doses of 8-3 mg/kg (one case) or 20 mg/kg plus 40 mg/kg (two cases) RU 486 by mouth (Mifepristone; Roussel-Uclaf, France) from day 26 to day 36 after the first day of mating of the bitch. Abortions occurred within two, four, 11 and 11 days after the initial treatment, respectively. The clinical status of the bitches was similar to that observed during a normal parturition, ie, lowering of the body temperature, shivering, panting and nesting behaviour. No side effects were seen. The beagle bitch that aborted twice, was mated at the first oestrus after the first abortion, conceived and aborted the same number of puppies the second time. The peripheral plasma progesterone concentration at the time of treatment in all bitches was < 75 nmol/litre. It had decreased to between 24-2 and 13-1 nmol/litre at the time of abortion and to between 4-0 to 0–5 nmol/litre at four to 15 days after the initial treatment. Peripheral plasma levels of prolactin increased three- to fourfold within 24 to 48 hours after treatment, concomitant with the drop in progesterone and had returned to basal levels within two to three days. Prolactin concentrations also increased around the time of intrauterine fetal death. Prostaglandin F_2aα-metabolite concentrations increased slowly after treatment, and around the time of abortion the levels increased five- to 10-fold. RU 486 seems to be a safe and effective abortifacient for use during mid-term pregnancy in the dog.     

Clinical signs and hormonal changes in dairy heifers after induction of parturition with prostaglandin F2 alpha

Parturition was induced in 12 dairy heifers with prostaglandin (PG) F2 alpha about 2 weeks before the expected time of calving. Eight animals gave birth after two injections (group 1), three animals needed more than two injections (group 2) and one animal (cow no. 740) required one injection. All animals in groups 1 and 2 had retained foetal membranes and the time needed to induce parturition was 59 +/- 7 and 149 +/- 10 h, respectively. As cow no. 740 did not have retained foetal membranes and calved 24 h after one PGF2 alpha injection, it was excluded from the results. Udder distension and relaxation of the pelvic ligaments could predict the calving to within 12 h. Furthermore, the pre-calving drop of body temperature could predict the time of parturition to within 16 h. The total white blood cells and polymorphonuclear cells were at their highest values on the day preceding parturition whereas mononuclear cells had a tendency to increase 3 days after calving. Increased levels of haemoglobin were found at the time of parturition, whereas, plasma-calcium levels significantly decreased after parturition (P < 0.001). Progesterone levels markedly decreased after the first PGF2 alpha injection and reached 2 nmol/l at the time of parturition. Plasma levels of oestradiol-17 beta reached the peak at the time of parturition, whereas, the highest levels of the PGF2 alpha metabolite and cortisol were recorded 16 h after calving.     

Validation of a direct radioimmunoassay of melatonin in the blue fox

A direct radioimmunoassay procedure for the determination of melatonin in the blood of blue fox has been validated and applied.The assay required 50 μl of sample and standard, 100 ul of antiserum and 100 μl of (³H)melatonin. After overnight incubation at 4°C the antibody bound melatonin was separated from the free hormone with dextran-coated charcoal. Following centrifugation the antibody bound (³H)melatonin was determined in a beta scintillation counter.The antiserum bound 30–35 % of the (³H)melatonin at a final dilution of 1:36000. The non specific binding represented less than 5% of the total radioactivity in all assays. The lowest detectable amount of melatonin was 2.6 fmol/tube, corresponding to 52.5 pmol/1. The inter-assay coefficient of variation at 178 and 510 pmol/1 was 15.6 and 8.8%, respectively. The precision profile, calculated from a 10-replicate standard curve, showed that the coefficient of variation decreased from 43% at 84 pmol/1 to 15% at 336 pmol/1, and remainded at or below 10% for concentrations exceeding 670 pmol/1.Plasma was collected from 2 male blue foxes at about hourly intervals during a 24 h period in September and assayed for melatonin. Maximum (421 pmol/1) and minimum (97 pmol/1) concentrations of the hormone were inversely related to light intensity.     

Trace element-rich litter in soils: influence on biochemical properties related to the carbon cycle

Purpose  

The aim of this work was to study the effect of ‘trace element-rich litter’ on the properties of two reforested polluted soils of different pH values (acidic and neutral) in terms of (1) availability of trace elements and (2) chemical and biochemical properties of the soil at different pH. We hypothesized that this litter would affect several parameters related to the organic matter cycle in soils, depending on initial soil pH.     

Ultrastructural studies on malignant lymphoma in dogs, with the use of computer imaging

Malignant lymphoma is one among the most often diagnosed malignant tumors in dogs. In the course of the disease lymphatic glands become enlarged and infiltrations form in internal organs or skin. Studies on the ultrastructure of neoplastic lymphocytes were carried out on a formalin-fixed and parafin-embedded material with a Hulquist and Karlsson's method. Numerous aplastic cells with large nuclei, abundan eu- or heterochromatin and unstabilised nucleolus were observed on TEM slides. The cells differed clearly from normal lymphocytes with highly condesed heterochromatin and stabilised nucleoli.     

Variability of the Content of Soluble Non-Digestible Polysaccharides in Rye Inbred Lines

The content of soluble non-digestible, non-starch polysaccharides (SNDP) in grain of about 700 inbred lines of rye varied during three seasons (1985–1987) from 35 to 75 mg/g with the highest frequency between 55 and 75 mg/g. The SNDP level in popular Polish open pollinated cultivars was never below 60 mg/g. The results of reciprocal crosses proved lack of xenia effect on the SNDP content. The level of SNDP compounds was not correlated with either protein or thousand kernel weight. In spite of large environmental effect of SNDP and protein contents the coefficients of variation of these characters in S₃ sublines in each environment were below 7 % and the heritability in broad sense amounted up to 0.73 for SNDP and 0.61 for protein. A few lines have been selected with low SNDP content transferred to their progenies. In some of them protein content was as high as 18%; these might be good donors of high nutritive value.     

Effect of fractionated weaning on hormonal patterns and weaning to oestrus interval in primiparous sows

The effect of weaning the 4-5 heaviest piglets in the litter on day 33 of lactation and the remainder 2 days later (fractionated weaning) on plasma levels of prolactin, cortisol, oestradiol-17 beta (E2), progesterone (P4) and LH, as well as on the weaning to oestrus interval in primiparous sows was studied. Twelve crossbred sows were grouped into 6 pairs according to farrowing date and litter size. The litter of 1 sow in each pair (F) was weaned in 2 stages, and the other conventionally weaned at 35 days (C). Blood samples were collected via a permanent jugular vein catheter every 3 h from 9 a m to 9 p m daily throughout the experimental period, and intensively at 15 min intervals for 12 h on the day of first and final weaning and for 6 h on the day after each weaning. All sows were slaughtered following their first post-weaning oestrus and the reproductive organs were macroscopically examined. Lactational oestrus was not observed in any of the sows. Sows from 5 out of 6 pairs showed oestrus within 8 days of weaning and post-mortem examination showed normal ovulation. There was a tendency for the F sows to have a shorter weaning to oestrus interval, as compared with the C sows (5 of 6 pairs, 4.8 days v 5.6 days). The plasma levels of prolactin around weaning were not significantly different between the 2 groups. Within 6 h after final weaning, the prolactin concentrations decreased gradually from 7.6 and 8.7 to 1.6 and 1.7 microgram/l in the control and treatment groups, respectively. The plasma levels of cortisol, showing a diurnal rhythm (with the lowest level at 6 and/or 9 p m), did on no occasion differ between the 2 groups. On the day of final weaning, no diurnal rhythm was observed, with cortisol remaining high at 6 and 9 p m. The plasma levels of E2 and P4 were low until final weaning in both groups. After final weaning the E2 levels rose faster in the F sows than in the C sows, to 44.3 and 34.8 pmol/l, respectively, on day 2 (p less than 0.01). No significant differences in levels of plasma LH and the number of LH pulses were observed between the groups. After final weaning the average and base levels of LH and the number of LH pulse(s) increased significantly.     

Immunocytochemical identification of neoplastic cell clone in phase S of cell cycle in transplantable rat tumors

Wistar and Buffalo rats of both sexes, aged 4 months, were divided into three groups: I which was given an intramuscular injection of 3 x 10(6) cells of Morris hepatoma (Buffalo males), II--subcutaneous injection of 3 x 10(4) cells of mammary gland carcinoma (Wistar females), III--intraperitoneal injection of 3 x 10(4) cells of Yoshid sarcoma (Wistar males). The animals were killed: in group I--19, group II--13 and in group III--6 days after tumor transplantation. Twenty four hours before euthanasia the rats were given 5-brome-2'-deoxyuridine (BRd-U) at a dose of 50 mg/kg body mass. The control group consisted of animals with tumour. They were not treated with BRd-U. Immunocytochemical reaction was performed on the sections of tumors, using monoclonal anti-BRd-U clone BU-33, Sigma. Computer measurements of tumor cells were carried out. There was a high similarity in morphological parameters between two kinds of cancer, and clear differences between them and Yoshid sarcoma. The main difference was noted in a twofold increase in the quantity of synthesised DNA in the nuclei of sarcoma cells. Immunocytochemical identification of tumor cells in phase S of the cell cycle with the use of monoclonal anti-BRd-U antibody is a precise and quick method of estimation of their proliferative potential.