Endometrial hyperplasia and mineralization in zoo felids treated with melengestrol acetate contraceptives.

Melengestrol acetate (MGA) contraceptives are widely used in zoo felids to regulate fertility and may have deleterious effects on endometrial health. To determine whether MGA exposure was associated with endometrial disease, the genital tracts of 212 zoo felids (99 MGA treated and 113 control) representing 23 species were evaluated. Adenomatous and cystic hyperplasia were prevalent in both MGA-treated (85%) and control (61%) groups, and the risk of developing these lesions increased with age. Treatment with MGA further increased the risk of developing advanced hyperplasia regardless of dose, and treatment for >72 months significantly elevated that risk, whereas parous animals had a lower risk. Endometrial polyps, fibrosis, adenomyosis, and hydrometra occurred in both MGA-treated and control animals. MGA treatment was associated with an increased risk of hydrometra and mineralization but not of adenomyosis, polyps, or fibrosis after adjusting for advanced hyperplasia. Acute or chronic endometritis were associated with advanced hyperplasia but not with MGA treatment. These results indicate that proliferative and inflammatory endometrial lesions are common spontaneous diseases in zoo cats, and MGA contraceptives increase the risk of some diseases. The association of MGA with endometrial lesions that could impair fertility should be considered when using this contraceptive in genetically valuable felids.

The effects of melengestrol acetate on the ovaries of captive wild felids.

Melengestrol acetate (MGA) is the most widely used contraceptive in zoo felids, but the mechanism of contraception and the pathologic effects have not been investigated. For this study, the effects of MGA on folliculogenesis were assessed, and the association of MGA with ovarian lesions was evaluated. Comparisons were made among the histopathologic findings in the ovaries from 88 captive wild felids (representing 15 species) divided into three groups: 37 currently contracepted with MGA, eight previously exposed to MGA, and 43 never contracepted. Ninety-one percent of the felids evaluated had tertiary follicles, and no differences were noted between contracepted and uncontracepted cats. Some MGA-contracepted cats also had corpora lutea indicating recent ovulation. These results indicate that folliculogenesis not suppressed by current doses of MGA and ovulation occurred in some cats. Therefore, the contraceptive actions of MGA do not occur by suppressing folliculogenesis, and MGA-contracepted felids likely have endogenous estrogens that may confound progestin effects on the uterus. Cystic rete ovarii was the most common pathologic finding, but they were not more prevalent in MGA-contracepted cats. These findings indicate that MGA is not associated with ovarian disease, including ovarian cancer, in contrast to the uterine lesions noted in MGA-treated cats.

Nutritional restriction and acid-base balance in white-tailed deer.

We examined the effect of progressive nutritional restriction on acid-base balance in seven captive, adult white-tailed deer (Odocoileus virginianus) from 4 February to 5 May 1988 in north central Minnesota (USA). Metabolic acidosis was indicated by low mean blood pH (7.25 to 7.33) in deer throughout the study. Mean urinary pH values declined (P = 0.020) from a mean (+/- SE) baseline of 8.3 +/- 0.1 to 6.7 +/- 0.3 as restriction progressed. Acidemia and aciduria were associated with significant variations in mean blood CO2 (P = 0.006) and pO2 (P = 0.032), serum potassium (P = 0.004) concentrations, and with a significant (P = 0.104) handling date x group interaction in urinary potassium: creatinine values. Mean bicarbonate:carbonic acid ratios were consistently below 20:1 during nutritional restriction. Mean packed cell volume increased (P = 0.019) and serum total protein decreased (P = 0.001); thus there was evidence for progressive dehydration and net protein catabolism, respectively. Blood pCO2, serum sodium, and urinary sodium:creatinine were stable throughout the study. We propose that acidosis and aciduria are metabolic complications associated with nutritional restriction of white-tailed deer.

Ability of thawed tiger (Panthera tigris) spermatozoa to fertilize conspecific eggs and bind and penetrate domestic cat eggs in vitro.

Electroejaculates from tigers were collected and half was used fresh to inseminate tiger eggs in vitro and domestic cat eggs stored in a hypertonic salt solution. The remainder was pelleted, frozen in a solution of 20% egg yolk, 11% lactose and 4% glycerol, thawed and cultured with tiger and domestic cat eggs. The motility index ((sperm % motility)+(status rating x 20))/2 for thawed spermatozoa was about 86% of that in fresh aliquots. Of the 49 tiger oocytes inseminated in vitro with fresh spermatozoa, 34 (69.4%) cleaved, compared with 33 of 47 oocytes (70.2%) cultured with thawed spermatozoa (P > 0.05). Embryos generated by either sperm treatment could develop in vitro to the 16-cell or morula stage. Fresh and thawed tiger spermatozoa were equally capable (P > 0.05) of binding and penetrating the outer and inner zona pellucida of domestic cat eggs. These results demonstrate the ability of frozen-thawed tiger spermatozoa to (i) penetrate homologous and heterologous eggs and (ii) result in conspecific, advanced development of preimplantation embryos in vitro.

Influence of hypothalamic-pituitary-adrenocortical hormones on reproductive hormones in gray wolves (Canis lupus).

The release of hypothalamic-pituitary-adrenocortical hormones was studied in intact and neutered gray wolves (Canis lupus) to determine how these hormones interact and affect reproductive hormones. Experiments were performed on adult wolves anesthetized with 400 mg ketamine and 50 mg promazine. Intravenous (i.v.) injections with 50 micrograms ovine corticotropin releasing factor (oCRF) significantly increased adrenocorticotropin (ACTH; P < or = 0.01), cortisol (CORT; P < or = 0.004), and progesterone (P < or = 0.036), but not beta-endorphin (P > or = 0.52). Since neutered wolves demonstrated dose-dependent elevations in response to ACTH, it was concluded that the progesterone was secreted from the adrenal gland. Basal luteinizing hormone (LH) concentrations in neutered wolves were similar before and 60 min after i.v. injection of 1, 5, or 25 IU ACTH (P > or = 0.36) or 2.2 mg/kg cortisol (P = 0.42). Neither 25 IU ACTH (P = 0.55) nor 0.22 mg/kg dexamethasone (P = 0.49) altered the LH response to injection of LH releasing hormone in neutered wolves. Chronic administration of 0.22 mg/kg/day dexamethasone for 3 d did not alter baseline LH concentrations (P = 0.75). Injection of 1.0 mg/kg naloxone (NAL), however, increased LH concentrations relative to baseline values in both intact (P = 0.032) and neutered (P = 0.0005) female wolves, but not in intact (P = 0.19) or neutered males (P = 0.07). These results indicated that in gray wolves (1) oCRF stimulated the release of pituitary and adrenal hormones in a fashion similar to that of other mammals; (2) the adrenal cortex was capable of secreting progesterone into the systemic circulation; (3) exogenous glucocorticoids did not alter LH concentrations; and (4) endogenous opioids may modulate LH secretion in female wolves.

Circannual prolactin rhythm in intact dogs housed outdoors.

Prolactin (PRL) and progesterone (P4) values were collected from eight intact (4 female, 4 male) mixed-breed dogs housed outdoors for a two-year span. A circannual component was significant for PRL for each dog (P less than 0.01) and the rhythm was validated for the population (P less than 0.004). Females had two estrus periods a year demonstrated by a P4 frequency of 23.5 +/- 0.47 wk. A 6-month component was statistically significant for P4 for three dogs and of borderline statistical significance for the fourth dog. Circannual acrophases yielded a statistically significant population rhythm (P = 0.012), whereas the 6-month component was of borderline statistical significance (P = 0.056). No time-macroscopic relationship between PRL cycles and P4 cycles was seen. These findings are consistent with similar PRL rhythms in non-domestic canids and emphasize the importance of considering seasonal effects when interpreting PRL data.

Gray wolf density and its association with weights and hematology of pups from 1970 to 1988.

We examined weights and hematologic profiles of gray wolf (Canis lupus) pups and the associated wolf density in the east-central Superior National Forest of northeastern Minnesota (USA) during 1970 to 1988. We collected weight and hematologic data from 117 pups (57 females, 60 males) during 1 September to 22 November each year. The wolf density (wolves/800 km2) trend was divided into three phases: high (72 +/- 7), 1970 to 1975; medium (44 +/- 2), 1976 to 1983; and low (27 +/- 2), 1984 to 1988. Wolf numbers declined (P = 0.0001) 39 and 63% from 1970 to 1975 to 1976 to 1983 and from 1970 to 1975 to 1984 to 1988, respectively. Weight was similar between male and female pups and did not vary as wolf density changed. Mean hemoglobin (P = 0.04), red (P = 0.0001) and white blood cells (P = 0.002), mean corpuscular volume, mean corpuscular hemoglobin concentration and mean corpuscular hemoglobin (P = 0.0001) did differ among the multi-annual phases of changing wolf density. Weight and hematologic data also were compared to values from captive wolf pups. The high, but declining wolf density was associated with macrocytic, normochromic anemia in wolf pups, whereas the lowest density coincided with a hypochromic anemia. Although hematologic values show promise for assessing wolf pup condition and wolf population status, they must be used cautiously until data are available from other populations.

Weights and hematology of wild black bears during hibernation.

We compared weights and hematological profiles of adult (greater than 3-yr-old) female black bears (Ursus americanus) during hibernation (after 8 January). We handled 28 bears one to four times (total of 47) over 4 yr of varying mast and berry production. Mean weight of lactating bears was greater (P less than 0.0001) than that of non-lactating females. White blood cells (P less than 0.05) and mean corpuscular volume (P = 0.005) also differed between lactating and non-lactating bears. Hemoglobin (P = 0.006) and mean corpuscular hemoglobin concentration (P = 0.02) varied among years; values were lowest during 1975, following decreased precipitation and the occurrence of a second year of mast and berry crop shortages in a three-year period. Significant (P less than 0.05) interaction between reproductive status (lactating versus non-lactating) and study year for hemoglobin, red blood cells, and packed cell volume, and increased mean corpuscular volume, suggested a greater nutritional challenge for lactating females compared to non-lactating females during the 1975 denning season. Our data suggest that hematological characteristics of denning bears may be more sensitive than weights as indicators of annual changes in nutritional status; however, other influential factors, in addition to mast and berry crop production, remain to be examined.

Differences in urinary chemistry profiles of moose on Isle Royale during winter.

During winters 1987-1988 (I) and 1988-1989 (II), we assessed the physiological status of moose (Alces alces) residing on the west and east ends of Isle Royale (Michigan, USA) by collecting and chemically analyzing urine deposited in snow (snow-urine) from January to early March. Samples were assayed for urea nitrogen (U), sodium (Na), potassium (K), calcium (Ca), phosphorus (P) and creatinine (C). Throughout both winters, elevated urinary U:C ratios in snow-urine samples collected from east-end moose compared to west-end moose indicated greater dietary energy deprivation and accelerated net catabolism of endogenous protein. Sodium: C ratios were low throughout the study and were similar between moose from both ends of the island, except during the middle of winter I. Greater K:C, P:C and Ca:C ratios in east-end moose compared to west-end moose throughout winter I, and increases in these ratios and U:C in east-end moose from middle to late winter during the second year provided additional evidence of a greater deterioration in condition in east-end moose. The superior nutrition provided to moose on the west end of the island was associated with more developed soils and diverse vegetation and a lower stem density of balsam fir compared to the east end.

Diazepam-induced feeding in captive gray wolves (Canis lupus).

Diazepam doses of 0.2, 0.4, and 0.8 mg/kg induced feeding in sated gray wolves in a dose-dependent manner (p less than 0.001). Neither 0.8 mg/kg of the benzodiazepine antagonist, beta-CCP (p = 0.36), nor 0.8 mg/kg of the benzodiazepine inverse agonist, beta-CCE (p = 0.85), decreased the diazepam-induced hyperphagia. Five of 6 naive wolves (p = 0.003) ate dry dog food within 15.4 +/- 1.9 min of being given 0.4 mg/kg diazepam and freely chose dog food after the single diazepam administration.

The influence of social and endocrine factors on urine-marking by captive wolves (Canis lupus).

Although serum hormones varied seasonally in all adult animals, only dominant male and female wolves urine-marked. Serum testosterone and urine-marking rates, which increased during the fall/winter breeding season, were positively correlated in both male and female dominant wolves. Estradiol, which increased in conjunction with proestrus and estrus, was not correlated with female urine-marking. These findings suggest that hormonal influence on urine-marking in the wolf is modulated by social factors and contrast with those for both domestic dogs and coyotes, two other members of the genus Canis.

In vitro fertilization and embryo development in vitro and in vivo in the tiger (Panthera tigris).

A study was conducted to evaluate the adaptability to the tiger of an in vitro fertilization/embryo culture system previously developed in the domestic cat. In Trial I (July 1989), 10 female tigers were treated with either 2,500 (n = 5) or 5,000 (n = 5) IU eCG i.m. and with 2,000 IU hCG i.m. 84 h later. In Trial II (January 1990), 6 females (5 of which were treated in Trial I) were given 2,500 IU eCG i.m. and 2,000 IU hCG i.m. 84 h later. Twenty-four to twenty-six hours after hCG treatment, all tigers were subjected to laparoscopy, and oocytes were aspirated transabdominally. On the basis of follicular development (follicles greater than or equal to 2 mm in diameter), all females responded to exogenous gonadotropins (range, 6-52 follicles/female). Follicle number and oocyte recovery rate were unaffected (p greater than 0.05) by eCG dose or time of year. A total of 456 oocytes were collected from 468 follicles (97.4% recovery; mean, 28.5 +/- 3.4 oocytes/female). Of these, 378 (82.9%) qualified as mature, 48 (10.5%) as immature, and 30 (6.6%) as degenerate. During Trial I, 8 electroejaculates were collected from 7 male tigers, and in Trial II, 3 semen samples were collected from 3 males. Motile sperm were recovered on each occasion; the overall mean (+/- SEM) ejaculate volume was 7.5 +/- 0.7 ml, the number of motile sperm/ejaculate was 105.9 +/- 20.6 x 10(6), and the percentage of structurally normal sperm/ejaculate was 81.4 +/- 2.0%. After swim-up processing, 0.05 x 10(6) motile sperm were co-cultured with 10 or fewer tiger oocytes in a humidified atmosphere (38 degrees C) of 5% CO2 in air. Of the 358 mature oocytes inseminated, 227 (63.4%) were fertilized. Oocytes from 2 females became contaminated in culture and, therefore, were excluded from embryo cleavage calculations. Of the remaining 195 fertilized oocytes, 187 (95.9%) cleaved to the two-cell stage. No parthenogenetic cleavage was observed in noninseminated control oocytes (n = 20). Eighty-six good-to-excellent-quality two- to four-cell embryos were transferred surgically into the oviducts of 4 of the original oocyte donors in Trial I and 2 females in Trial II. A pregnancy occurred in 1 female in Trial II, and 3 live-born cubs were delivered by Caesarean section 107 days after embryo transfer. Of the 56 cleaved embryos cultured in vitro in Ham's F10 for 72 h, 14 (25.0%) were at the sixteen-cell stage, and 15 (26.8%) were morulae.(ABSTRACT TRUNCATED AT 400 WORDS)

Immobilization of gray wolves (Canis lupus) with sufentanil citrate.

Gray wolves (Canis lupus) were immobilized with 0.5 mg/kg xylazine plus 7.5 micrograms/kg of either sufentanil (n = 8), etorphine (n = 8), or carfentanil (n = 2). Drug doses used in this study were selected to provide consistency for comparison and are not recommended doses for effective immobilization of wolves. Induction times were similar among groups (11.9 +/- 1.0 min). Thirty min after induction, wolves were given either 0.5 mg/kg naloxone hydrochloride plus 0.15 mg/kg yohimbine hydrochloride or saline only intravenously. Arousal times for wolves given naloxone and yohimbine (1.2 +/- 0.1 min) were shorter than wolves given saline (35.5 +/- 6.4 min). Respiratory rates were similar among the three drug groups (6.9 +/- 1.0 breaths/min). One animal given sufentanil then saline was found dead 108 min after induction. Presumptive diagnosis was renarcotization and hypothermia. Results indicated that sufentanil is an effective opioid immobilizing agent for gray wolves.

Effect of season on seminal traits and serum hormone concentrations in captive male Siberian tigers (Panthera tigris).

Electroejaculates and serum samples were collected throughout the year from 5 male Siberian tigers. Semen quality, seminal plasma chemistry and serum hormone concentrations were evaluated and analysed as a function of season. Semen volume, concentration, motility, viability and morphology were not influenced by season. The ability of spermatozoa to undergo capacitation and penetrate zona-free hamster eggs did not differ due to season. Season had no influence on seminal plasma concentrations of acid phosphatase, calcium, glucose, potassium and phosphorus. Serum values of thyroxine and triiodothyronine were lowest (P = 0.002 and P = 0.03, respectively) during the summer months while cortisol showed no seasonal variation. Serum testosterone concentrations were higher in fall and early winter (P less than 0.001) than at other times of the year but were not correlated with semen quality. These results demonstrate that season has no effect on the semen production and quality of captive male Siberian tigers.

Physiological responses of red foxes (Vulpes vulpes) to surgery.

Radio transmitters were surgically implanted into the abdomens of red foxes (Vulpes vulpes). Blood samples were taken before, immediately after, and 8 hr after surgery and analyzed for hormonal, biochemical, electrolyte and hematologic changes. Samples were taken at the same times from control foxes. Adrenocorticotropin increased after surgery (P less than 0.05), but returned to pre-surgery values after 8 hr. Cortisol increased and remained elevated in the surgery group relative to pre-surgery values or to control values (P less than 0.05); Triiodothyronine and thyroxine both decreased from post-surgery values 8 hr later (P less than 0.05). Creatine kinase, total bilirubin and aspartate aminotransferase increased after 8 hr in both surgery and control groups (P less than 0.05). Carbon dioxide increased under anesthesia in both groups, but returned to initial values after 8 hr (P less than 0.05). The white blood cell count increased after 8 hr only in the surgery group (P less than 0.05). There were no differences between the groups for any value obtained from the initial blood sample. These data indicate that abdominal surgery results in prolonged adrenocortical activity and decreased thyroid hormone levels, but otherwise has minimal systemic effects in red foxes.

Blood and urinary profiles of free-ranging desert mule deer in Arizona.

As a corollary to a more comprehensive study on their ecology, we documented blood and urinary profiles for 10 free-ranging desert mule deer (Odocoileus hemionus crooki) (five males, five females) captured by net-gun shot from a helicopter during February 1988 in Saguaro National Monument, Arizona. Pursuit with the helicopter for netting deer ranged from 3 to 15 min. Blood profiles included seven hematological characteristics and 12 serum chemistries, electrolytes, hormones and enzymes. Urine samples were assayed for urea nitrogen, creatinine, sodium, potassium, calcium and phosphorus. Urinary data were compared as ratios to creatinine. Serum cholesterol was greater (P less than 0.05) in males than females. Pursuit time was correlated with serum non-esterified fatty acids (r = 0.67, P less than 0.05) and influenced urinary specific gravity (r2 = 0.77, P less than 0.004), urea nitrogen: creatinine (r2 = 0.79, P less than 0.005), and potassium: creatinine (r2 = 0.42, P = 0.08) ratios. Increasing specific gravity was related to urinary creatinine concentration (r2 = 0.72, P less than 0.008). All deer exhibited acute adrenal stimulation, accompanied by elevated serum creatine phosphokinase and urinary potassium: creatinine ratios, which were indicative of acute excitement and muscle trauma associated with the capture process. We demonstrated that urinary data are a valuable supplement to serum data in demonstrating effects of intense physical exertion, and both forms of data emphasize the need to assess capture-related excitability as a source of variation in blood and urine characteristics of free-ranging desert mule deer.

Physiological and behavioral responses of gray wolves (Canis lupus) to immobilization with tiletamine and zolazepam.

We conducted a series of experiments to examine the efficacy of Telazol (TEL) for immobilization of captive gray wolves (Canis lupus). Ten wolves were immobilized with either 5 or 10 mg/kg TEL. There was no difference in induction time (6.5 +/- 0.8 versus 5.8 +/- 1.2 min; P = 0.63) between the two doses, but the time to initial arousal was longer for the higher dose (P = 0.0008). Wolves were again immobilized with 10 mg/kg TEL and upon initial arousal were given additional doses of either 5.0 mg/kg TEL or 2.5 mg/kg ketamine (KET) to maintain immobilization. Wolves given boosters of TEL had longer second recovery times than wolves given KET (P = 0.01). There were no differences in induction times or arousal times for wolves immobilized with TEL that had been reconstituted with sterile water and stored at 20 C for 30 days (P greater than or equal to 0.11) or 60 days (P greater than or equal to 0.27) when compared to immobilization times using fresh solution. Induction times for wolves given TEL reconstituted with water and propylene glycol and stored for 60 days at -9 C were longer (P less than 0.05) than such times for wolves given standard TEL, but time to initial arousal was unchanged (P greater than or equal to 0.44). There were no differences in heart rates (P = 0.36), blood pressures (P = 0.32), respiratory rates (P = 0.91), and rectal temperatures (P = 0.62) between the two TEL doses. Telazol was shown to be an effective and safe immobilizing agent for gray wolves.

Chemical immobilization of red foxes (Vulpes vulpes).

Red foxes (Vulpes vulpes) were immobilized with one of the following drug combinations: ketamine/xylazine (n = 22), ketamine/promazine (n = 35), ketamine/midazolam (n = 13), or tiletamine/zolazepam (n = 22). Foxes given ketamine/xylazine had the shortest induction and longest recovery times relative to other drug combinations, whereas foxes given ketamine/midazolam had the longest induction times. Recommended doses for the various combinations are given. Foxes given ketamine/xylazine were given either 0.1, 0.2, 0.4 mg/kg yohimbine, or saline 40 min after anesthetic induction. Administration of yohimbine significantly shortened arousal and recovery times relative to control values (P less than 0.001).

Fertility control in female white-tailed deer.

Silastic rods containing either melengestrol acetate (MGA) or levonorgestrel (LN) were placed in anestrous white-tailed deer (Odocoileus virginianus borealis) does to evaluate the contraceptive efficacy of the implants over a 2 yr period. Implants of MGA were placed in five does during mid-pregnancy to evaluate the effect of this treatment on pregnancy, parturition and lactation. Pregnancies were not observed in the five animals implanted with MGA during anestrus. Three of five does implanted with LN became pregnant in the first season. Pregnancy was not interrupted in the five pregnant does implanted with MGA and it was necessary to remove the implants and treat the does with an estrogen to achieve parturition. One of five fawns was delivered alive and was raised by the doe. MGA was effective for 2 yr as a contraceptive in white-tailed deer, LN was ineffective as used, and MGA placed in pregnant does delayed or prevented normal parturition and thus should not be used in pregnant deer.

In-vitro induction of capacitation of fresh and frozen spermatozoa of the Siberian tiger (Panthera tigris).

Electroejaculates from 5 tigers were split and half of each was assayed fresh while the remainder was frozen and thawed before being assayed. Preincubation time, temperature and removal of seminal plasma were evaluated for their effect on in-vitro capacitation. Ability of spermatozoa to penetrate oocytes, as measured by the zona-free hamster egg-sperm penetration assay (SPA), was used as verification of capacitation. Results of the experiments with fresh semen indicate that: (1) preincubation time affects the fertilizability of tiger spermatozoa with 2 h appearing optimal, (2) a preincubation temperature of 37 degrees C results in significantly higher penetration rates than does a 22 degrees C treatment, and (3) tiger seminal plasma does not appear to contain decapacitation factors, as has been reported for several other species. Frozen semen experiments indicate that (1) frozen-thawed tiger spermatozoa must be removed from the environment of the semen extender before capacitation can take place, and (2) the freeze-thaw procedure results in a shortening of the required capacitation time.