Hypoglycemic activity of CPU2206: A novel peptide from sika (Cervus nippon Temminck) antler.

Previous work had extracted and purified an antidiabetic peptide named CPU2206 with 7,127.6 Da. In this work, the toxicity of CPU2206 was first evaluated by daily administration to ICR mice, and after 28 days of administration, the body weight and lipid metabolism of the mice did not change significantly, which proved its safety and reliability. Second, further studies have focused on its hypoglycemic effects by daily intraperitoneal injection to alloxan-induced diabetic mice and KK-Ay mice, showing that CPU2206 effectively decreased the blood glucose and corresponding indicators of diabetic mice. Daily administration of CPU2206 nearly normalized the lipid metabolic parameters in diabetic mice. Histological examination also validated that CPU2206 ameliorated the pancreas injuries induced by alloxan or alleviated islet hypertrophy caused by insulin resistance in KK-Ay mice. To sum up, a totally new bioactive peptide CPU2206 obtained from sika antler showed significantly antidiabetic as well as lipid-lowering effects in diabetic mice. PRACTICAL APPLICATIONS: Antler has been used as a traditional Chinese medicine to invigorate primordial energy, enrich the blood, strengthen bones, and improve both male and female sexual functions for thousands of years. Traditionally, velvet antler can be grinded directly and taken orally, or used in porridge, wine and meat stew. Our experiment enriches the research on the function of edible antlers, provides the basis for developing it into functional health food, and on the other hand, provides an idea for finding new antidiabetic drugs.

WNT4 acts downstream of BMP2 to mediate the regulation of ATRA signaling on RUNX1 expression: Implications for terminal differentiation of antler chondrocytes.

Although ATRA is involved in regulating the proliferation and differentiation of chondrocytes, its underlying mechanism remains unknown. Here we showed that ATRA could stimulate the proliferation of antler chondrocytes and expression of COL X and MMP13 which were two well-known markers for hypertrophic chondrocytes. Silencing of CRABP2 prevented the induction of ATRA on chondrocyte terminal differentiation, while overexpression of CRABP2 exhibited the opposite effects. CYP26A1 and CYP26B1 weakened the sensitivity of antler chondrocytes to ATRA. Further analysis evidenced that ATRA might induce chondrocyte terminal differentiation and modulate the expression of BMP2, WNT4, and RUNX1 through RARα/RXRα. Knockdown of BMP2 enhanced the induction of ATRA on the expression of COL X and MMP13, whereas overexpression of BMP2 abrogated this effectiveness. WNT4 might mediate the effects of ATRA and BMP2 on chondrocyte terminal differentiation. Dysregulation of BMP2 impaired the regulation of ATRA on WNT4 expression. Administration of ATRA to antler chondrocytes transfected with RUNX1 siRNA failed to induce the differentiation. Conversely, rRUNX1 strengthened the stimulation of ATRA on the expression of COL X and MMP13. Simultaneously, RUNX1 was a downstream effector of BMP2 and WNT4 in chondrocyte terminal differentiation. Moreover, WNT4 might play an important role in the crosstalk between BMP2 and RUNX1. Attenuation of BMP2 or WNT4 enhanced the interaction between ATRA and RUNX1, while constitutive expression of BMP2 or WNT4 reversed the regulation of ATRA on RUNX1. Collectively, WNT4 may act downstream of BMP2 to mediate the effects of ATRA on the terminal differentiation of antler chondrocytes through targeting RUNX1.

Does Cu supplementation affect the mechanical and structural properties and mineral content of red deer antler bone tissue?

The main factors affecting the mechanical (and other) properties of bone, including antler, are the proportions of ash (especially Ca and P) and collagen content. However, some trace minerals may also play more important roles than would be expected, given their low levels in bone and antler. One such trace mineral is Cu. Here, we studied the effects of Cu supplementation on the mechanical and structural characteristics, and mineral content of antlers from yearling and adult (4 years of age) red deer fed a balanced diet. Deer (n=35) of different ages (21 yearlings and 14 adults) were studied. A total of 18 stags (11 yearlings and 7 adults) were injected with Cu (0.83 mg Cu/kg BW) every 42 days, whereas the remaining 17 (10 yearlings and 7 adults) were injected with physiological saline solution (control group). The Cu content of serum was analysed at the beginning of the trial and 84 days after the first injection to assess whether the injected Cu was mobilized in blood. Also, the mechanical and structural properties of antlers and the mineral content in their cortical walls were examined at three (yearlings) or four (adults) points along the antler beam. The effect of Cu supplementation was different in yearlings and adults. In yearlings, supplementation increased the Cu content of serum by 28%, but did not affect antler properties. However, in adults, Cu supplementation increased the Cu content of serum by 38% and tended to increase the cortical thickness of antlers (P=0.06). Therefore, we conclude that, even in animals receiving balanced diets, supplementation with Cu could increase antler cortical thickness in adult deer, although not in yearlings. This may improve the trophy value of antlers, as well as having potential implications for bones in elderly humans, should Cu supplementation have similar effects on bones as those observed in antlers.

CONTROLLING ANTLER GROWTH IN A CASTRATED INDOCHINESE SIKA DEER CERVUS NIPPON PSEUDAXIS USING A COMMERCIALLY AVAILABLE TRENBOLONE ACETATE AND ESTRADIOL IMPLANT.

A captive Indochinese sika deer (Cervus nippon pseudaxis) was castrated at the age of 5 yr. The resultant abnormal antler growth over the next few years became difficult to manage from both the veterinary and husbandry standpoints. Using a commercially available trenbolone acetate and estradiol implant marketed for domestic cattle heifers, normal mineralization of the abnormal antlers was achieved along with the expected normal casting. The deer was then maintained for 6 yr using an annual implant regimen.

Effect of ruminally protected methionine on body weight gain and growth of antlers in red deer (Cervus elaphus) in the humid tropics.

Sixteen male red deer (Cervus elaphus), with an initial weight of 93.83 ± 4.38 kg were used to study the effect of the dietary level of ruminally protected methionine (RPM) on growth, antler morphometry and serum metabolites. The deer were allowed to graze and were individually supplemented with concentrate to provide 0, 2.5, 3.5 or 4.5 g/day of RPM for 65 days. There was a quadratic effect on daily weight gain (0.163, 0.098, 0.147 and 0.172 kg/day; P < 0.005) and antler beam length (59.7, 63.6, 63.0 and 57.2 cm; P < 0.02) for RPM levels of 0 to 4.5 g/day, respectively. The brow tine length and the number of points were similar across all treatments. Serum glutamic pyruvate transaminase (SGPT) was reduced (quadratic effect; P < 0.02; 22.0, 29.6, 18.0 and 17.8 IU/l) to RPM levels of 0 to 4.5 g/day, respectively, whereas other metabolites (glucose, cholesterol, urea, creatinine, total protein and serum glutamic oxaloacetic transaminase [SGOT]) were unchanged. Increasing the RPM dose did not affect weight gain or antler growth. Methionine supplementation reduced the activity of SGPT.

Testis and antler dysgenesis in sitka black-tailed deer on Kodiak Island, Alaska: Sequela of environmental endocrine disruption?

It had been observed that many male Sitka black-tailed deer (Odocoileus hemionus sitkensis) on Kodiak Island, Alaska, had abnormal antlers, were cryptorchid, and presented no evidence of hypospadias. We sought to better understand the problem and investigated 171 male deer for phenotypic aberrations and 12 for detailed testicular histopathology. For the low-lying Aliulik Peninsula (AP), 61 of 94 deer were bilateral cryptorchids (BCOs); 70% of these had abnormal antlers. Elsewhere on the Kodiak Archipelago, only 5 of 65 deer were BCOs. All 11 abdominal testes examined had no spermatogenesis but contained abnormalities including carcinoma in situ-like cells, possible precursors of seminoma; Sertoli cell, Leydig cell, and stromal cell tumors; carcinoma and adenoma of rete testis; and microlithiasis or calcifications. Cysts also were evident within the excurrent ducts. Two of 10 scrotal testes contained similar abnormalities, although spermatogenesis was ongoing. We cannot rule out that these abnormalities are linked sequelae of a mutation(s) in a founder animal, followed by transmission over many years and causing high prevalence only on the AP. However, based on lesions observed, we hypothesize that it is more likely that this testis-antler dysgenesis resulted from continuing exposure of pregnant females to an estrogenic environmental agent(s), thereby transforming testicular cells, affecting development of primordial antler pedicles, and blocking transabdominal descent of fetal testes. A browse (e.g., kelp) favored by deer in this locale might carry the putative estrogenic agent(s).

The effect of cyproterone acetate on the antler cycle in red deer (Cervus elaphus L.).

The aim of the study was to test the effect of antiandrogen, cyproterone acetate (CA) on the antler cycle in the red deer (Cervus elaphus). CA was administered to three adult red deer stags (Edward, Fuks and Gacek) in weekly intervals. Edward and Fuks were given 600 mg + 600 mg of CA, whereas Gacek was given 600 mg + 300 mg. CA was injected during the hard antler phase: in mid-October (Edward), at the end of November (Fuks) and at the end of January (Gacek). CA caused the antler casting 17 to 22 days after the first injection. In all stags, the casting of antlers was followed by a period of intensive growth of new antlers. Edward was given CA at the end of October. This treatment was responsible for occurrence of the two antler cycles in the year of the experiment. When CA was administered during the middle of the hard antler phase an additional short antler cycle occurs followed by new antler growth. CA treatment in the later part of hard antler phase may cause a prolonged antler cycle.

Effects of testosterone on pedicle formation and its transformation to antler in castrated male, freemartin and normal female red deer (Cervus elaphus).

Pedicles and antlers are male deer secondary sexual characters. As such, development of these structures is under the control of androgen hormones. Pedicle growth is caused by increasing and elevated plasma testosterone (T) levels, whereas first antler transformation from a fully formed pedicle occurs when the T levels are decreasing. Castration prior to pedicle initiation abrogates future pedicle and antler formation. Female deer also have the potential to develop pedicles and antlers, but they do not normally express this phenotype due to lack of sufficient androgen stimulation. Previous studies have shown that female white-tailed deer could be readily induced to grow pedicles as well as antlers by singular administration of exogenous androgens (EA), but in red deer (Cervus elaphus) singular or irregular EA treatment could only stimulate castrated male, normal or ovariectomised females to grow pedicles, but not antlers. The present study was set out to test whether these EA-induced pedicles in red deer failed to give rise to antlers was because they were constitutively incapable of doing so, or because the plasma T profile naturally exhibited in intact stags was not achieved by the androgen treatment used in these previous studies. Eight castrated red deer stag calves, 3 freemartins (females which were born co-twin to males), and 3 normal female red deer were used in the present study and treated with EA, either as biweekly injections for the castrates or as implants for freemartin and females until the late stage of pedicle growth. Blood sampling was carried out biweekly for the analyses of plasma T and IGF1 concentration. The results showed that the natural plasma T profile in the experimental deer was successfully mimicked through regular EA treatment and subsequent withdrawal at late pedicle growth stage. All castrated males, 2 out of 3 freemartin, and 1 out of 3 normal female red deer formed not only pedicles, but also antlers. Based on these results, we conclude that EA-induced pedicles at least in red deer of the genus Cervus, like those in the genus Odocoileus, are constitutively capable of giving rise to antlers, if they are of sufficient height.

Effect of zeranol or melengestrol acetate (MGA) on testicular and antler development and aggression in farmed fallow bucks.

Fifteen yearling fallow bucks were randomly assigned by BW to one of three treatment groups: control (C; n = 5), melengestrol acetate (MGA; n = 5), and zeranol (Z; n = 5), to evaluate effects on testicular development, aggressive behavior, antler growth, sexual activity, ADG, and BW. Zeranol-treated bucks received zeranol ear implants (36 mg) at 90-d intervals, and MGA-treated bucks received MGA in the ration (100 microg x animal(-1) x d(-1)). Bucks grazed ryegrass/Coastal bermudagrass pasture and were supplemented with 3:1 corn/soybean meal at 0.45 kg x animal(-1) x d(-1). Body weights, body condition scores (BCS), blood samples, and testis measurements were obtained at d 0 and at 14-d intervals for 229 d. As bucks reached hard antler (7/15 to 8/25), antlers were harvested and weighed, and ejaculates were collected at 14-d intervals. Aggression was evaluated using 10-min video sessions scoring body blows, avoidance, head pushes, and head bunts. Scrotal circumference (SC) and paired testis volume were affected by a day x treatment interaction (P < 0.01); testes of zeranol-treated bucks were smaller than those of control or MGA-treated bucks. First sperm in the ejaculate tended to be delayed (P < 0.10) in zeranol-treated bucks compared with control and MGA-treated bucks. Melengestrol acetate-treated bucks had a maximum sperm concentration in the ejaculate that was three times (P < 0.05) that of control bucks and nine times (P < 0.05) that of zeranol-treated bucks. Antler weight was the least (P < 0.01) for bucks receiving zeranol and greatest (P < 0.10) for MGA-treated bucks; intermediate values were recorded for the control bucks. Aggressive behavior was delayed (P < 0.05) for zeranol-treated bucks until treatment effects were overcome. Melengestrol acetate-treated bucks had decreased (P < 0.01) aggressive behavior compared with control bucks. Melengestrol acetate-treated bucks had increased (P < 0.05) serum testosterone concentrations compared with control and zeranol-treated bucks. Human chorionic gonadotropin-stimulated peak serum testosterone concentrations for zeranol-treated bucks were delayed (P < 0.01) compared with control and MGA-treated bucks. Although zeranol-treated bucks overcame treatment effects, they were never able to reach testicular measurements or sperm concentrations equal to those of the control or MGA-treated bucks. Zeranol and MGA treatments may have both positive and negative effects that can be utilized when producing slaughter bucks.

No direct mitogenic effect of sex hormones on antlerogenic cells detected in vitro.

Deer pedicles, antecedents of antlers, develop from a specialized periosteum (antlerogenic periosteum) which overlies the lateral crest of the deer frontal bone. The initiation of pedicle growth is triggered by androgen hormones. Thus far, it is not known whether pedicle initiation is caused by direct stimulation of androgen hormones on the antlerogenic periosteum or whether some intermediate mechanisms are necessary. The present study took an in vitro approach to investigate whether sex hormones have direct mitogenic effects on primary cultured antlerogenic periosteal cells (antlerogenic cells). Antlerogenic cells were obtained from two 5-month-old red deer calves. The cells were passaged twice and then treated with testosterone, dihydrotestosterone, and estradiol. The proliferation assays showed that no direct mitogenic effects on the second passage antlerogenic cells could be detected with any of the sex hormone treatments (P > 0.05). Testosterone-binding studies showed that at the second passage, specific testosterone-binding sites were present in the antlerogenic cells. Therefore, we conclude that androgens do not have mitogenic effects on antlerogenic cells in vitro. Our results suggest that pedicle formation may not be the result of direct stimulation of androgen hormones on antlerogenic tissue. Instead, androgen hormones may only allow the process to proceed by increasing the sensitivity of antlerogenic cells to mitogens, e.g., some growth factors.

Cyproterone acetate reduced antler growth in surgically castrated fallow deer.

We studied the role of androgens in antler growth. In particular, we investigated whether the onset of antler regrowth is triggered by a short-term pulse of testosterone and if low levels of androgens are required for antler growth. The study was conducted on 12 surgically castrated fallow deer bucks (Dama dama) aged approximately 27 months. Six animals (CA group) were given the antiandrogen, cyproterone acetate (CA, 1000 mg/treatment); the others were given vehicle solution only (control). Before each CA treatment, blood was sampled and analysed for testosterone, androstenedione, IGF-1, cortisol, FSH, and LH. CA treatment and blood sampling were performed 2 days before castration, on the day of castration and afterwards at 2-day intervals until day 22. Subsequently, CA treatment and blood sampling continued at weekly intervals until day 270. All animals cast their antlers, followed by antler regrowth in all control bucks, but in only four of the six CA-treated castrates. Plasma testosterone concentrations were low in all animals (between 0.01 and 0.20 ng/ml), but were significantly (P<0001) greater in the controls. In both groups, a temporary increase in testosterone values was recorded around the time of antler regrowth, the peak being significantly (P<0.01) higher in the controls. Androstenedione showed a similar pattern as testosterone. Plasma IGF-1 concentrations increased sharply during the antler growth spurt and did not differ significantly between the two groups throughout the study period. Cortisol concentrations were greater in controls than in the CA group. However, no link with the antler cycle was apparent. FSH and LH concentrations were higher in the controls for most of the study. Antlers produced by the control bucks were significantly larger than those in the CA group (P<0.03). For antler length, testosterone, androstenedione and IGF-1, areas under the curve (AUC) were calculated over the period of antler growth. For the pooled deer (n=12) significant correlations existed between AUCs of antler length and testosterone, but not for antler length and IGF-1. Also, a trend for a positive correlation between AUCs of antler length and androstenedione was noted. It is concluded that a plasma androgen concentration at least above a minimal threshold level is a necessary prerequisite for normal antler regrowth in fallow deer, and that this androgen effect is not mediated via circulating IGF-1. The biological role of low levels of androgens may be to sensitize antler cells to the stimulating effect of IGF.

Treatment of the growing pedicle with retinoic acid increased the size of the first antlers in fallow deer (Dama dama L.).

Unilateral injection of 10 mg of all-trans retinoic acid (RA) into the lateral portion of the growing pedicle of fallow bucks (n = 20) led to a significant (P = 0.033, Wilcoxon matched-pairs test) increase in first antler volume (median, 25.5 ml) as compared to the contralateral (control) side, injected with vehicle only (median, 21.5 ml). It is hypothesized that the RA treatment of the developing pedicle exercised a direct or indirect effect on the periosteal/perichondrial cells covering the growing cranial appendage, resulting in an increased proliferation rate of the cells of the antler perichondrium.

Effects of retinoic acid on pedicle and first antler growth in a fallow buck (Dama dama L.).

Unilateral injection of 10 mg all-trans retinoic acid (in 1 ml of castor oil) into the early pedicle anlage of a male fallow deer resulted in accelerated growth of the cranial appendage, and altered pedicle and first antler shape on the treatment side, whereas pedicle and antler growth on the control side, injected with 1 ml of vehicle solution only, was normal. It is concluded that retinoic acid is able to alter pedicle and first antler morphogenesis, presumably by affecting the positional information of the periosteal cells covering the incipient pedicle.

Role of steroids in antler growth of red deer stags.

A series of six studies were carried out in red deer stags to test hypotheses concerning the importance of steroid control of velvet antler growth and to investigate mechanisms by which these hormones exert their effects. Medroxyprogesterone acetate (MPA) an LH inhibitor administered to stags during hard antler caused premature antler casting, reduced subsequent antler weight and caused a reduction in the LH and testosterone responses to GnRH. In two separate studies blockade of testosterone receptors with cyproterone acetate (CPA) administered to stags, either during early velvet antler growth or during the hard antler stage, significantly reduced LH and testosterone responses to GnRH. In both studies antler length, but not weight, was increased by CPA treatment. In another study testosterone implants were used to prevent the gradual decline in plasma testosterone levels normally observed during winter. Implants were removed 3 weeks before the anticipated date of antler casting. The implants significantly increased plasma testosterone levels and subsequent antler growth (expressed as a proportional increase compared with the previous year) compared with untreated controls. To determine whether the annual cycle of plasma testosterone response following GnRH stimulation was due simply to a lack of LH stimulation, ovine LH was injected on six occasions at defined stages of the antler cycle to red deer stags and the testosterone response measured. The testosterone responses were low at antler casting and during velvet antler growth compared with antler cleaning and peak rut. It appears low testosterone levels are due, in part, to a loss of responsiveness by the testes to LH as well as a low level of secretion of LH during the antler growing season. Finally synthetic ACTH was injected at the same defined stages of antler growth as in the previous study to determine whether cortisol and adrenal androgen production altered with the stage of the antler cycle. No significant differences were found in the dehydroepiandrosterone (DHEA) response, but cortisol responses were higher from late velvet antler growth to peak rut, compared with the times of antler casting and early velvet growth. Overall it was concluded that velvet antler growth can occur without testosterone stimulation during the period of velvet growth, but the data reinforce the concept that the timing of antler growth is linked to the annual cycle of testosterone.

Effects of insulin-like growth factor-I (IGF-I) and IGF-II on the growth of antler cells in vitro.

The effects of insulin-like growth factors -I and -II (IGF-I and -II) on the growth of undifferentiated (fibroblast zone) cells from the growing tip of red deer velvet antlers and from cells 1.5 cm distal to the growing tip (cartilage zone) were investigated in primary cell culture. The addition of IGF-I or IGF-II to the medium of cultures preincubated in serum-free medium for 24 h increased the rate of [3H]thymidine uptake in a dose-dependent manner in both cell types, with maximal stimulation occurring when 1 nM-30 nM was added. The addition of IGF-II to the incubation medium containing IGF-I did not cause a further increase in [3H]thymidine uptake in either cell type over and above each growth factor alone, indicating that there were unlikely to be synergistic effects of IGF-II on the mitogenicity of IGF-I. Binding studies were carried out using 3 x 10(5) fibroblast zone cells and cartilage zone cells after they had been incubated in serum-free medium for 24 h. 125I-Labelled IGF-I (10(-9) M) in a final volume of 200 microliters was added to each culture and incubation carried out at 4 degrees C for a further hour. 125I-Labelled IGF-I bound specifically to both fibroblasts and cartilage zone cells; binding was displaced by both unlabelled IGF-I and by IGF-I antibody.(ABSTRACT TRUNCATED AT 250 WORDS)

Cells cultured from the growing tip of red deer antler express alkaline phosphatase and proliferate in response to insulin-like growth factor-I.

Deer antler growth provides a unique natural model of rapid and complete bone regeneration. In this study, the distal antler tips of male red deer (Cervus elaphus) were collected post-mortem during the annual growth period (April-August), and an in vitro system established for the culture of cells from three regions; the inner layer of the perichondrium, the reserve mesenchyme and the cartilage zone. Alkaline phosphatase (ALP) expression by cultured cells, as demonstrated by enzyme histochemistry and biochemical assay, reflected the stage of cellular differentiation. ALP activity was highest in cells cultured from the hypertrophic cartilage region (3.6 +/- 0.2 mumol/micrograms cell protein/minute), and lowest in undifferentiated mesenchymal cells (0.3 +/- 0.01 mumol/microgram cell protein/minute). ALP expression was lost with passage in culture. Levels of ALP activity in cultured cells correlated with the pattern and extent of enzyme expression in tissue sections as demonstrated by histochemical staining. Insulin-like growth factor (IGF)-I (10(-9)M-10(-7)M) was found to be mitogenic for cultured cells from all three zones as shown by increased incorporation of [3H]thymidine into DNA. These results demonstrate that cells from three different regions of the antler tip can be maintained in culture, and that antler cells share certain phenotypic characteristics of growth plate chondrocytes. These data provide further evidence of a role for IGF-1 in the regulation of antler growth. Antler regrowth is a potentially useful model for the study of the factors that regulate bone formation.

Effects of an antiandrogen treatment on morphological characters and physiological functions of male fallow deer (Dama dama L.).

From mid-April to late November, i.e., during antler growth phase and main breeding season, a group of four (later three) male fallow deer was treated with increasing doses of the antiandrogen cyproterone acetate (CA). Throughout the treatment period, plasma testosterone levels of the bucks were either undetectable or only slightly above the detection limit of 0.1 ng/ml. In marked contrast to controls, testicular volumes and neck girths of the CA treated bucks were basal during the application period, proving the strong antigonadotropic effect of CA (testicular volume) and the absence of androgen action at the normal target organs (neck girths), respectively. Antler growth in the CA treated bucks occurred during the normal time span for fallow deer and the antlers produced were of species specific shape. From this it is concluded that in fallow deer, onset, duration, and termination of antler growth are independent of androgen action. Velvet shedding in the CA treated bucks however did not occur at the normal time (late August/early September), but was postponed until late December/early January, i.e., about 5 weeks after the termination of CA application. Activity of alkaline phosphatase in the experimental animals increased during the antler growth phase, but was markedly lower than in controls. Maximum body weight in the CA treated bucks was reached already in June, i.e., about 3 months prior to controls. Furthermore, the rut associated, drastic decline in body weight observed in controls did not occur in the experimental animals.

Effects of an antiandrogen treatment on the antler cycle of male fallow deer (Dama dama L.).

Weekly application of the antiandrogen cyproteron acetate (CA) to 3 adult (group 1, treatment starting 3 weeks prior to the rut) and 4 subadult (group 2, treatment starting after the rut) fallow bucks led to premature antler casting after about 9 weeks (group 1) or 16 to 17 days (group 2). Variation in time span between onset of CA administration and induced casting (in late November/early December) is attributed to age- and season-dependent differences in androgen levels before and during the treatment period. Casting surfaces of the antlers were flat. In all bucks, casting was followed by a complete though shortened antler cycle, occurring under short day conditions. In February/March velvet was shed from the antlers, indicating a rise in plasma androgen levels at that time. At time of fraying, antlers exhibited signs of immaturity and were shorter than normal. In the most pronounced case of immaturity, the antlers consisted mainly of cancellous woven bone, revealing that bone remodeling and antler mineralization had not occurred to a larger extent. These observations prove that onset of fraying behaviour does not depend on prior maturation of antler bone or necrosis of velvet. Occurrence of the additional antler cycle in late autumn and winter had no negative effect on subsequent antler formation starting in the following spring.

Rapid neural growth: calcitonin gene-related peptide and substance P-containing nerves attain exceptional growth rates in regenerating deer antler.

Deer antler is a unique mineralized tissue which can produce very high growth rates of > 1 cm/day in large species. On completion of antler growth, the dermal tissues which cover the antler are shed and the underlying calcified tissue dies. After several months the old antler is discarded and growth of a new one begins. It is known that deer antlers are sensitive to touch and are innervated. The major aims of this study were to identify and localize by immunohistochemical techniques the type of innervation present, and to find out whether nerve fibres could exhibit growth rates comparable to those of antler. We have taken tissue sections from the tip and shaft of growing Red deer (Cervus elaphus) antlers at three stages of development; shortly after the initiation of regrowth, the rapid growth phase, and near the end of growth. Incubation of tissue sections with antisera to protein gene product 9.5 (a neural cytoplasmic protein), neurofilament triplet proteins (a neural cytoskeletal protein), substance P and calcitonin gene-related peptide (both of which are present in and synthesized by sensory neurons) showed the presence of immunoreactive nerve fibres in dermal, deep connective and perichondrial/periosteal tissues at all stages of antler growth. The sparse distribution of vasoactive intestinal polypeptide-like immunoreactivity was found in dermal tissue only at the earliest stage of antler development. Nerve fibres immunoreactive to neuropeptide Y, C-flanking peptide of neuropeptide Y and tyrosine hydroxylase, all present in postganglionic sympathetic nerves, were not observed at any stage of antler growth. Nerves expressing immunoreactivity for any of the neural markers or peptides employed could not be found in cartilage, osteoid or bone. These results show that antlers are innervated mainly by sensory nerves and that nerves can attain the exceptionally high growth rates found in regenerating antler.

Reproduction in the red deer female and the effect of oestrogens on the antler cycle and behaviour.

Red deer (Cervus elaphus) is a seasonally breeding mammal. The season of reproduction is probably regulated by photoperiod. The rut and the majority of conceptions occur in October. If hind is not allowed to mate the oestrus is repeated at an interval of about 13.5 days. In captivity the mating season is more prolonged and the calving season is extended. The length of gestation is about 235 days. The fat % of milk varies between 6.6 and 17.4. A hind in good condition may attain puberty as 16.5 month old. The smaller doses of stilboestrolum dipropionicum (S, 150-250 mg) induced mainly a female type of sexual behaviour or oestrus. The bigger and more prolonged administration of S (250-300 mg, given twice with a monthly interval) induced a strong male sexual behaviour e.g. roaring, chasing a hind in oestrus, mouting a hind in oestrus or a dummy sprinkled with pheromones and performance of an ejaculatory peak (M). During M the hind was in an almost vertical position and a single ejaculation of saliva was always observed. The artificially induced antlers of hinds without hormonal treatment became broken and their regrowth was very small. After S the velvet was always shed. After larger doses of S the casting was more delayed and the next antler growth was bigger.