ESLAV/ECLAM/LAVA/EVERI recommendations for the roles, responsibilities and training of the laboratory animal veterinarian and the designated veterinarian under Directive 2010/63/EU.

Directive 2010/63/EU was adopted in September 2010 by the European Parliament and Council, and became effective in January 2013. It replaces Directive 86/609/EEC and introduces new requirements for the protection of animals used for scientific purposes. In particular, it requires that establishments that breed, supply or use laboratory animals have a designated veterinarian (DV) with expertise in laboratory animal medicine, or a suitably qualified expert where more appropriate, charged with advisory duties in relation to the well-being and treatment of the animals. This paper is a report of an ESLAV/ECLAM/LAVA/EVERI working group that provides professional guidance on the role and postgraduate training of laboratory animal veterinarians (LAVs), who may be working as DVs under Directive 2010/63/EU. It is also aimed at advising employers, regulators and other persons working under the Directive on the role of the DV. The role and responsibilities of the DV include the development, implementation and continuing review of an adequate programme for veterinary care at establishments breeding and/or using animals for scientific purposes. The programme should be tailored to the needs of the establishment and based on the Directive's requirements, other legislations, and current guidelines in laboratory animal medicine. Postgraduate laboratory animal veterinary training should include a basic task-specific training module for DVs to complement veterinary competences from graduation, and continuing professional development on the basis of a gap analysis. A tiered approach to further training in laboratory animal veterinary medicine and science offers career development pathways that are mutually beneficial to LAVs and establishments.

Evidence for a major gene affecting the transition from normoglycaemia to hyperglycaemia in Psammomys obesus.

We investigated the mode of inheritance of nutritionally induced diabetes in the desert gerbil Psammomys obesus (sand rat), following transfer from low-energy (LE) to high-energy (HE) diet which induces hyperglycaemia. Psammomys selected for high or low blood glucose level were used as two parental lines. A first backcross generation (BC(1)) was formed by crossing F(1) males with females of the diabetes-prone line. The resulting 232 BC(1) progeny were assessed for blood glucose. All progeny were weaned at 3 weeks of age (week 0), and their weekly assessment of blood glucose levels proceeded until week 9 after weaning, with all progeny maintained on HE diet. At weeks 1 to 9 post weaning, a clear bimodal distribution statistically different from unimodal distribution of blood glucose was observed, normoglycaemic and hyperglycaemic at a 1:1 ratio. This ratio is expected at the first backcross generation for traits controlled by a single dominant gene. From week 0 (prior to the transfer to HE diet) till week 8, the hyperglycaemic individuals were significantly heavier (4--17%) than the normoglycaemic ones. The bimodal blood glucose distribution in BC(1) generation, with about equal frequencies in each mode, strongly suggests that a single major gene affects the transition from normo- to hyperglycaemia. The wide range of blood glucose values among the hyperglycaemic individuals (180 to 500 mg/dl) indicates that several genes and environmental factors influence the extent of hyperglycaemia. The diabetes-resistant allele appears to be dominant; the estimate for dominance ratio is 0.97.

Elevated hypothalamic beacon gene expression in Psammomys obesus prone to develop obesity and type 2 diabetes.

OBJECTIVE: To investigate hypothalamic beacon gene expression at various developmental stages in genetically selected diabetes-resistant and diabetes-prone Psammomys obesus. In addition, effects of dietary energy composition on beacon gene expression were investigated in diabetes-prone P. obesus. METHODS: Hypothalamic beacon gene expression was measured using Taqman fluorogenic PCR in 4-, 8- and 16-week-old animals from each genetically selected line. RESULTS: Expression of beacon was elevated in the diabetes-prone compared with diabetes-resistant P. obesus at 4 weeks of age despite no difference in body weight between the groups. At 8 weeks of age, hypothalamic beacon gene expression was elevated in diabetes-prone animals fed a high-energy diet, and was correlated with serum insulin concentration. CONCLUSION: P. obesus with a genetic predisposition for the development of obesity and type 2 diabetes have elevated hypothalamic beacon gene expression at an early age. Overexpression of beacon may contribute to the development of obesity and insulin resistance in these animals.

Psammomys obesus and the albino rat--two different models of nutritional insulin resistance, representing two different types of human populations.

Animal models for insulin resistance and type 2 diabetes are required for the study of the mechanism of these phenomena and for a better understanding of diabetes complications in human populations. Type 2 diabetes is a syndrome that affects 5-10% of the adult population. Hyperinsulinaemia, hypertriglyceridaemia, decreased high-density lipoprotein (HDL) cholesterol levels, obesity and hypertension, all form a cluster of risk factors that increase the risk of coronary artery disease, and are known as insulin resistance syndrome or syndrome X. The gerbil, Psammomys obesus is characterized by primary insulin resistance and is a well-defined model for dietary induced type 2 diabetes. Weanling Psammomys and Albino rats were held individually for several weeks on high energy (HE) and low energy (LE) diets in order to determine the development of metabolic changes leading to diabetes. Feeding Psammomys on HE diet resulted in hyperglycaemia (303 +/- 40 mg/dl), hyperinsulinaemia (194 +/- 31 microU/ml) and a moderate elevation in body weight, obesity and plasma triglycerides. Albino rats on HE diet demonstrated an elevation in plasma insulin (30 +/- 4 microU/ml), hypertriglyceridaemia (170 +/- 11 mg/dl), an elevation in body weight and obesity, but maintained normoglycaemia (98 +/- 6 mg/dl). Psammomys represent a model that is similar to human populations, with primary insulin resistance expressed in young age, which leads to a high percentage of adult type 2 diabetes. Examples for such populations are the Pima Indians, Australian Aborigines and many other Third World populations. The results indicate that the metabolism of Psammomys is well adapted towards life in a low energy environment, where Psammomys takes advantage of its capacity for a constant accumulation of adipose tissue that will serve for maintenance and breeding in periods of scarcity. This metabolism known as 'thrifty metabolism', is compromised at a high nutrient intake.

Circulating lipoproteins and hepatic sterol metabolism in Psammomys obesus prone to obesity, hyperglycemia and hyperinsulinemia.

The liver plays a central role in lipoprotein metabolism and cholesterol homeostasis. As the physiopathology of lipid disorders in non-insulin-dependent diabetes mellitus (NIDDM) is multifactorial and still imperfectly known, we evaluated its onset on plasma lipid transport and hepatic cholesterol metabolism in Psammomys obesus. This sand rat lapses into hyperinsulinemia and hyperglycemia when transferred from its native food to laboratory rodent diets. Marked hypertriglyceridemia and hypercholesterolemia developed in hyperinsulinemic (Group B) and hyperglycemic/ hyperinsulinemic (Group C), compared with normal P. obesus (Group A). Group B showed significantly (P<0.05) higher plasma VLDL-cholesterol (41.9%) and LDL-cholesterol (47.3%) concentrations, whereas Group C was characterized by an even more marked increase in VLDL-cholesterol (176%, P<0.001) compared with Group A. Lipoprotein composition was also altered, displaying impaired lipid and apolipoprotein moiety distribution in IDL, LDL, HDL(2) and HDL(3) lipoprotein fractions of Groups B and C. The activity of hepatic 3-hydroxy-3-methylglutaryl coenzyme A reductase, the rate-limiting enzyme in cholesterol biosynthesis, was consistently lower in Group B (P<63.4%, P<0.001) and C (43.9%, P<0.005). In contrast, the direct measurement of microsomal acyl-CoA:cholesterol acyltransferase (ACAT), controlling the acylation of cholesterol, showed an increase averaging 53% in Group B (P<0.01) and 61% in Group C (P<0.005). Similarly, elevated activity (171.1%, P<0.05 and 291.4%, P<0.001, respectively) was related to cholesterol 7alpha-hydroxylase, the rate-limiting enzyme in bile acid biosynthesis. These alterations were accompanied with abundant deposition of triglycerides and cholesterol in the liver. Changes in circulating lipids and liver parameters were related to glucose and insulin levels, indicating the implication of insulin resistance and diabetes. Therefore, our findings demonstrate various disturbances in plasma lipid profile and lipoprotein composition, as well as in liver cholesterol metabolism during the sequential development of insulin resistance and diabetes in P. obesus rats. Furthermore, the current data point to an undoubtedly important role of the liver in the pathogenesis of metabolic disorders in the progression of nutritionally-induced insulin resistance and diabetes in P. obesus. Finally, current research shows that more marked plasma and hepatic lipid perturbations occur in insulin resistance than in diabetes, which may culminate in the development of atherosclerosis.

The role of person familiarity in young infants' perception of emotional expressions.

This research investigated the role of person familiarity in the ability of 3.5-month-old infants to recognize emotional expressions. Infants (N = 72) were presented simultaneously with two filmed facial expressions, happy and sad, accompanied by a single vocal expression that was concordant with one of the two facial expressions. Infants' looking preferences and facial expressions were coded. Results indicated that when the emotional expressions were portrayed by each infant's own mother, infants looked significantly longer toward the facial expressions that were accompanied by affectively matching vocal expressions. Infants who were presented with emotional expressions of an unfamiliar woman did not. Even when a brief delay was inserted between the presentation of facial and vocal expressions, infants who were presented with emotional expressions of their own mothers looked longer at the facial expression that was sound specified, indicating that some factor other than temporal synchrony guided their looking preferences. When infants viewed the films of their own mothers, they were more interactive and expressed more positive and less negative affect. Moreover, infants produced a greater number of full and bright smiles when the sound-specified emotion was "happy," and particularly when they viewed the happy expressions of their own mothers. The average duration of negative affect was significantly longer for infants who observed the unfamiliar woman than for those who observed their own mothers. These results show that when more contextual information-that is, person familiarity-was available, infants as young as 3.5 months of age recognized happy and sad expressions. These findings suggest that in the early stages of development, infants are sensitive to contextual information that potentially facilitates some of the meaning of others' emotional expressions.

Changing pattern of prevalence of insulin resistance in Psammomys obesus, a model of nutritionally induced type 2 diabetes.

Psammomys obesus (a desert gerbil, nicknamed the "sand rat") with innate insulin resistance was transferred to a high-energy (HE) diet at a young (8 to 20 weeks) and older (38 to 45 weeks) age. The young Psammomys progressed to in vivo insulin resistance, followed by pronounced hyperglycemia and hyperinsulinemia, as described previously. Analysis of the time dependency of these changes in response to the HE diet showed that the increase in serum glucose preceded the increase in insulin and plateaued earlier, reverting to normal together with insulin in the older Psammomys. Implants releasing insulin 2 IU/24 h did not induce appreciable hypoglycemia, a decrease in free fatty acids (FFAs), or a suppression of hepatic phosphoenolpyruvate carboxykinase (PEPCK) activity in young animals after 5 hours, despite a markedly increased circulating insulin. However, in the older Psammomys, the exogenous hyperinsulinemia produced a significant decline in serum glucose and FFA and a suppression of hepatic PEPCK activity. A euglycemic-hyperinsulinemic clamp confirmed that hepatic glucose production (HGP) was lower in older Psammomys versus the young and was almost completely abolished by insulin (from 5.6 +/- 0.6 to 0.2 +/- 0.1 mg x min(-1) x kg(-1) v 10.9 +/- 0.8 to 3.9 +/- 0.5 mg x min(-1) x kg(-1)). This indicates that HGP, rather than glucose underutilization, was the main contributor to the hyperglycemia and that the hepatic insulin resistance in Psammomys is attenuated with age. In relation to the human condition, these findings point out that while the type 2 diabetes prevalence in Western populations generally increases with age, the excessive nutritional intake in high-risk populations produces a pattern of diabetes prevalence that tapers off with age. As such, the nutritionally induced diabetes in Psammomys represents a similar model for a differing pattern of the age-related prevalence of diabetes.

Role of renal nerves in hemodynamic and natriuretic responses to saline in rabbits with impaired baroreflex sensitivity.

The hemodynamic and urinary Na+ excretory response to a 2.5-fold increase in NaCl by i.v. infusion were assessed in conscious male rabbits with either high (BShi, salt-insensitive) or low (BSlo, salt-sensitive) cardiac baroreflex sensitivity, before, and 11-14 days after bilateral renal denervation. Effective renal plasma flow (ERPF) and proximal tubular Na+ reabsorption were measured by para-amino-hippurate (PAH) and Li+ clearances, respectively, before and after NaCl infused for 2 hr at a rate of 0.11 mL/kg/min. Intact BShi rabbits, showed a significant natriuresis within 30 min which was associated with an increase in ERPF and inhibition of proximal tubular reabsorption. The Na+ excretion rate was much slower in BSlo rabbits, while ERPF and proximal tubular reabsorption remained unchanged. Renal denervation reduced MAP, increased basal ERPF, Na+ and Li+ excretion in both groups, and abolished the difference in the renal hemodynamic re-sponse and Li+ excretion to increased NaCl, but not that in the rate of Na+ excretion. The data suggest that BSlo rabbits do not increase their ERPF and Li+ in response to saline because of an inability to bring about an inhibition of renal sympathetic nerve activity. This could be due to an impairment in the sensitivity of their cardiopulmonary baroreceptors. The difference in the rate of natriuresis in the two groups of rabbits which remained after renal denervation could involve an additional hormonal or a local renal mechanism.

Predicting variation in the timing of language milestones in the second year: an events history approach.

In a longitudinal investigation of 40 child-mother dyads, we examined prediction from three indexes of children's own language: (1) vocal imitations, (2) first spontaneous words in production, and (3) receptive language starting at 0;9, and their mothers' verbal responsiveness at 0;9 and 1;1, to the developmental onset of three significant language milestones of the second year: (1) 50 words in productive language, (2) combinatorial speech, and (3) the use of language to express a memory. In these analyses, we utilized EVENTS HISTORY ANALYSIS, a statistical technique well suited to questions concerning when in development certain events begin and the extent to which predictors influence the timing of those events. The timing of children's first words in production, the timing of their achievement of 50 words in receptive language, and maternal responsiveness at 1;1 each contributed uniquely to variation in the timing of the three language milestones. When child and mother factors were considered together, the onset of the three language milestones differed by as much as 0;5 months for children in the lower and upper 10th percentiles of the predictor variables. The present findings contribute to generating and testing specific models about child and mother factors thought to explain variation in key aspects of children's second-year language development.

Not all (possibly) "random" sequences are created equal.

The need to assess the randomness of a single sequence, especially a finite sequence, is ubiquitous, yet is unaddressed by axiomatic probability theory. Here, we assess randomness via approximate entropy (ApEn), a computable measure of sequential irregularity, applicable to single sequences of both (even very short) finite and infinite length. We indicate the novelty and facility of the multidimensional viewpoint taken by ApEn, in contrast to classical measures. Furthermore and notably, for finite length, finite state sequences, one can identify maximally irregular sequences, and then apply ApEn to quantify the extent to which given sequences differ from maximal irregularity, via a set of deficit (def(m)) functions. The utility of these def(m) functions which we show allows one to considerably refine the notions of probabilistic independence and normality, is featured in several studies, including (i) digits of e, pi, radical2, and radical3, both in base 2 and in base 10, and (ii) sequences given by fractional parts of multiples of irrationals. We prove companion analytic results, which also feature in a discussion of the role and validity of the almost sure properties from axiomatic probability theory insofar as they apply to specified sequences and sets of sequences (in the physical world). We conclude by relating the present results and perspective to both previous and subsequent studies.

Insulin resistance in the NIDDM model Psammomys obesus in the normoglycaemic, normoinsulinaemic state.

The desert gerbil Psammomys obesus ("sand rat"), a model of nutritionally induced insulin resistance and non-insulin-dependent diabetes mellitus, was treated after weaning with exogenous insulin implants in the normoglycaemic, normoinsulinaemic state. Albino rats matched for weight and age served as high energy diet adjusted reference animals. Insulin administration, elevating the serum insulin to 6000 pmol/l resulted in only a mild reduction in blood glucose levels in Psammomys, but caused a severe, often fatal hypoglycaemia in the albino rats. The hepatic response to insulin-induced hypoglycaemia in rats involved a significant loss in glycogen and suppression of phosphoenolpyruvate carboxykinase (PEPCK) activity. In Psammomys under similar hyperinsulinaemia no appreciable changes in liver glycogen and PEPCK activity were evident, indicating that blood glucose was replenished by continuing gluconeogenesis. Euglycaemic, hyperinsulinaemic clamp caused a complete shut-down of hepatic glucose production in albino rats. However, in both diabetes-prone and diabetes-resistant Psammomys lines, mean hepatic glucose production was reduced by only 62 to 53% respectively, despite longer lasting and higher levels of hyperinsulinaemia. These results indicate that Psammomys is characterized by muscle and liver insulin resistance prior to diet-induced hyperglycaemia and hyperinsulinaemia. This is assumed to be a species feature of Psammomys, exemplifying a metabolic adjustment to survival in conditions of food scarcity of both animal and human populations. It may reflect a propensity to insulin resistance and hyperglycaemia in population groups exposed to affluent nutrition.

Renal denervation prevents sodium retention and hypertension in salt-sensitive rabbits with genetic baroreflex impairment.

1. Rabbits with a genetic impairment in baroreflex control of heart rate become hypertensive on a high salt diet. The present study determined the effect of bilateral renal denervation on blood pressure and sodium balance after salt loading (four times normal intake; 28-36 mEq NaCl/day) in normotensive rabbits with high (Group I) and low (Group II) baroreflex sensitivity, respectively. 2. Eight rabbits in each group were denervated or sham-denervated 1 week before commencement of the high salt diet. Before operation, the two groups differed only in the gain of their cardiac baroreflex (Group I, -6.4 +/- 0.4 beats min-1 mmHg-1; Group II, -3.2 +/- 0.15 beats min-1 mmHg-1). 3. In Group I sham-denervated rabbits, mean arterial pressure remained unchanged, and plasma renin activity and heart rate fell significantly in response to the high salt. In Group II sham-denervated rabbits, mean arterial pressure increased by 10.6 +/- 1.2 mmHg, and heart rate and plasma renin activity remained unchanged. Their cumulative Na+ retention and weight gain was more than twice that of Group I sham-denervated rabbits. 4. Renal denervation decreased plasma renin activity in both groups to < 1 pmol Ang I h-1 ml-1, lowered cumulative Na+ retention from 102 +/- 4 to 35 +/- 5 mEq (P < 0.01) and completely prevented the increase in mean arterial pressure in response to high salt in Group II. 5. The results suggest that Group II rabbits retain salt and fluid in response to their diet because of an abnormality in their control of renal nerve activity, possibly via vagal afferents. This results in blood pressure elevation because of an inability to lower peripheral resistance and heart rate in response to the increase in cardiac output. 6. Since they display several of the characteristics of salt-sensitive hypertensive humans, i.e. salt retention, normal plasma renin activity, but abnormal regulation of plasma renin activity and blood flow in response to salt loading, Group II are an appropriate model of human salt-induced hypertension.

Immunocytochemical investigation of insulin secretion by pancreatic beta-cells in control and diabetic Psammomys obesus.

Hyperproinsulinemia is a characteristic feature of non-insulin-dependent diabetes mellitus (NIDDM) caused by pancreatic beta-cell dysfunction through a secretion-related alteration or impaired proinsulin processing. We have investigated the insulin processing and secretion in Psammomys obesus fed with low- and high-energy diets, which represent a model for diet-induced NIDDM. With a high-energy diet the animals develop hyperglycemia and hyperinsulinemia, whereas those maintained on a low-energy diet remain normoglycemic. Although a large amount of insulin immunoreactivity was detected in beta-cells of the normoglycemic compared to hyperglycemic animals, in situ hybridization for insulin mRNA demonstrated a particularly high signal in the beta-cells of the hyperglycemic animals. By electron microscopy, the beta-cells of normoglycemic animals displayed large accumulations of secretory granules, whereas those of the hyperglycemic animals contained very few granules and large deposits of glycogen. These results reflect a secretory resting condition for the cells of the normoglycemic animals in contrast to stimulated synthetic and secretory activities in the cells of the hyperglycemic ones. Using colloidal gold immunocytochemistry at the electron microscopic level, we have examined subcellular proinsulin processing in relation to the convertases PC1 and PC2. Immunolabeling of proinsulin, insulin, C-peptide, PC1, and PC2 in different cell compartments involved in beta-cell secretion were evaluated. Both PC1 and PC2 antigenic sites were detected in beta-cells of hyperglycemic Psammomys, but their labeling intensity was weak compared to the cells of normoglycemic animals. In both groups of animals, higher levels of PC2 were found in the Golgi apparatus than in the immature granules. Major decreases in proinsulin, insulin, PC1, and PC2 immunoreactivity were recorded in beta-cells of the hyperglycemic Psammomys. In addition, all these antigenic sites were detected in lysosome-like structures, revealing a major degradation process. These results suggest that the insulin-secreting cells in hyperglycemic Psammomys obesus are in a chronic secretory state during which impaired processing of proinsulin appears to take place.

Compressing data volume of left ventricular cineangiograms.

In this paper a method of compressing data volume for left ventricular cineangiograms is proposed. This method enables digital-optical discs to store the cineangiograms with an equivalent recording density to that of HDTV VTR's while preserving the quality of the original image. The data volume of the cineangiograms is compressed by approximating function and storing its coefficients. With this method, each cineangiogram frame is first decomposed into three regions: the inner part, the inner wall, and the background (using a statistical method previously reported by the authors). Each region is then compressed by means of a difference operation and an adaptive approximation with smooth functions. Performance is tested on 20 cases using actual cineangiograms. The specifications were verified for a spatial resolution of 1000 TV lines, a dynamic range of 60 dB, an SNR of 40 dB [pp/rms], volume compression to 7% of the original volume, and 0.19 [second/frame] for decoding on a 1.7 MFLOPS computer.

The efficiency of sand rat metabolism is responsible for development of obesity and diabetes.

Two separate lines--diabetic and partially diabetes-resistant--have been isolated from the sand rat (Psammomys obesus), each with different growth characteristics in response to diets of varying digestible caloric densities (high energy, HE, 2.93 kcal/g, or low energy, LE, 2.38 kcal/g). Over a two week period all animals consumed similar quantities (c. 125 g) irrespective of the diet consumed. Weight gains were as follows: diabetic line on HE diet - 59.7 g, on LE - 46.2 g; non-diabetic animals from the diabetes-resistant line on HE - 44 g. Only animals from the diabetic line, fed the HE diet, developed hyperinsulinemia, obesity and diabetes. The energy cost of weight gain for the diabetic line fed either HE or LE diets was 6.0 - 6.3 kcal/g whereas for the diabetes-resistant line on the HE diet, the cost of growth was 50% higher at 9.3 kcal/g. These differences could be due either to alterations in the content of tissue laid down or to differences in energy expenditure. It has already been established that diet-induced obesity and diabetes develop in the diabetic line with features typical of insulin resistance in the metabolism of the pancreas, liver and peripheral tissues. Some of the animals of the diabetes-resistant line may also develop diabetes over a long time period and go through a phase of transient hyperinsulinemia-normoglycemia. This may represent an intermediate stage in the development of the diabetic syndrome and serve as a model of type 2 diabetes in man.

Insulin dependence of murine T-cell lymphoma. II. Insulin-deficient diabetic mice and mice fed low-energy diet develop resistance to lymphoma growth.

Physiological concentrations of insulin support the in vitro growth of LB T-cell lymphoma. We could not detect similar insulin dependence in other tumor cell lines. This study reports that insulin also enhances the growth of LB cells in vivo. Mice treated with Streptozotocin (SZ) developed partial resistance to LB lymphoma growth and they survived longer (p < 0.0025) than non-diabetic mice after LB-cell inoculation. A few diabetic mice developed complete tumor resistance, manifested by total regression of the lymphoma. SZ-treated diabetic mice reconstituted with external insulin died as fast as non-diabetic mice when both were inoculated with the same number of LB cells. The SZ-treated diabetic mice did not develop resistance to the growth of BCLI B-cell leukemia, which demonstrated only a marginal proliferative response to insulin in vitro. Mice fed a low-energy diet exhibited low insulin levels and also developed resistance to lymphoma growth (50% survival 21 days vs. 15 days; p < 0.0005), supporting the concept that insulin enhances LB T-cell tumor development in mice.

Cyclohexyladenosine reduces MPTP induced tremor in dogs.

Male dogs were injected i.v. with 1 methyl-4-phenyl-1,2,3,6-tetrahydro- pyridine (MPTP) 2.5 mg/kg twice with an interval of 6 days. Twenty minutes after MPTP injection cyclohexyladenosine (CHA) 1 mg/kg i.v. was injected. Ten minutes after CHA injection the MTPT induced tremor, tachycardia and salviation ceased.