I. Rozenboim

Pattern of Pax7 Expression During Myogenesis in the Posthatch Chicken Establishes a Model for Satellite Cell Differentiation and Renewal

The paired‐box transcription factor Pax7 plays a critical role in the specification of satellite cells in mouse skeletal muscle. In the present study, the position and number of Pax7‐expressing cells found in muscles of growing and adult chickens confirm the presence of this protein in avian satellite cells. The expression pattern of Pax7 protein, along with the muscle regulatory proteins MyoD and myogenin, was additionally elucidated in myogenic cultures and in whole muscle from posthatch chickens. In cultures progressing from proliferation to differentiation, the expression of Pax7 in MyoD+ cells declined as the cells began expressing myogenin, suggesting Pax7 as an early marker for proliferating myoblasts. At all time points, some Pax7+ cells were negative for MyoD, resembling the reserve cell phenotype. Clonal analysis of muscle cell preparations demonstrated that single progenitors can give rise to both differentiating and reserve cells. In muscle tissues, Pax7 protein expression was the strongest by 1 day posthatch, declining on days 3 and 6 to a similar level. In contrast, myogenin expression peaked on day 3 and then dramatically declined. This finding was accompanied by a robust growth in fiber diameter between day 3 and 6. The distinctions in Pax7 and myogenin expression patterns, both in culture and in vivo, indicate that while some of the myoblasts differentiate and fuse into myofibers during early stages of posthatch growth, others retain their reserve cell capacity. Developmental Dynamics 231:489–502, 2004.

Various light source treatments affect body and skeletal muscle growth by affecting skeletal muscle satellite cell proliferation in broilers

In this study we addressed the effect of various monochromatic light treatments on muscle growth and satellite cell proliferation in broilers (Gallus domesticus). Broilers were reared under green (560 nm), blue (480 nm) and red (660 nm) monochromatic lights and white light as a control from day one until 35 days of age. At five days of age, satellite cells were prepared from the experimental chicks. The number of satellite cells per gram of breast muscle and total number of satellite cells derived from the experimental broilers was substantially higher in the groups reared under green and blue light, compared to the red and white light groups. Growth hormone receptor gene expression was also higher in the former groups. High correlation was found between the breast muscle weight observed on day 35 and the number of satellite cells per gram of breast muscle (r = 0.915) and total number of satellite cells (r = 0.833), derived from the experimental chicks as early as five days of age. In addition, the protein/DNA ratio found in breast muscle at 35 days of age was significantly lower in chicks that were reared under green and blue lights. The lowest ratio which was found in the green group and was twice as low as in the control group, indicates the highest number of nuclei in the former group. As satellite cells are the only source of additional nuclei in skeletal muscles of postnatal animals, our results suggest that the higher muscle weight found in the green and blue light groups was due to increased satellite cell proliferation during the first days of age.

Effect of light source and regimen on growing broilers

The aim of this study was to determine the effect of different light sources and light schedules on the growth and quality of commercial broilers. In each experiment 810 broiler chicks were divided into 3 groups, 3 replicates per group. All were reared at 20 lux. Body weight and food consumption were recorded weekly. Experiment 1. Birds were reared under 3 light sources: incandescent light bulb, warm-white fluorescent light tube or warm-white mini-fluorescent light bulb. Experiment 2. Birds were reared on 3 light schedules. 23 h light and 1 h dark (23L: 1D) throughout; an increasing light schedule with initial 23L:1D then 8L: 16D increasing daylight gradually to 16L:8D or an intermittently increasing daylight schedule (16:8P) where light and dark periods were shorter but portioned to achieve the same total hours per day up to 16L:8D. Broilers reared under mini-fluorescent light bulb were heavier than those under fluorescent tubes or incandescent bulbs by 49 d. Until 42 d of age, photoperiod had no effect on growth. However, at 49 d broilers reared under 16:8P and 16L:8D regimens were heavier than those or 23L:1D. At 42 d, female broilers on 23L:1D, were heavier than those on 16L:8D and 16:8P. Mortality was higher in groups on 23L:1D than on 16L:8D on 16:8P. At 49 d incidence of leg condemnation was higher in the 16:8P group. However, skin damage was lower in this group than in those on 23L: 1D and 16L:8D.

Enhancement of meat production by environmental manipulations in embryo and young broilers

Enhancing meat production by genetic selection for growth has already produced 5-week-old broilers weighing more than 2 kg. As growth performance characteristics continue to improve and the time it takes to achieve market size decreases, the period of embryonic development becomes a greater proportion of the birds life. Therefore, in parallel to genetic selection, other approaches, such as environmental manipulations in the embryo or in the early days posthatch, are becoming more relevant for increasing muscle growth and meat production. Recently, we have shown that nutritional treatments, i.e., providing feed immediately posthatch, or environmental treatments, such as heat conditioning or monochromatic green-light illumination during the first days posthatch, increase muscle growth and breast muscle weight at marketing day. In all cases, the increase in muscle growth was due to changes at the cellular and molecular levels leading to increased satellite cell proliferation and differentiation. The significant effects on muscle growth resulting from the treatments in the first days posthatch raised the hypothesis that muscle growth could be affected during the embryonic development. In experiments in which eggs were illuminated under monochromatic green light from embryonic day 5 (E5), there was a positive effect on embryo development and posthatch muscle growth. Further studies revealed that this enhanced muscle weight was due to increased satellite cell number and fiber synchronization during early days posthatch. Thermal manipulation at 38.5°C from E16 to E18 for 3 h/day had a delayed effect on satellite-cell proliferation and differentiation, resulting in enhanced hypertrophy of myofibers at market age.

Vitamin A deficiency interferes with proliferation and maturation of cells in the chicken small intestine

1. The effect of vitamin A on the small intestine was examined in vitamin-A-deficient meat-type chickens. 2. Maturation and activity of the small intestinal cells were assayed by detection of proliferating cells with proliferating cells nuclear antigen, goblet cells with Alcian blue, mature cells with alkaline phosphatase and extent of RNA expression with dot blot analysis. 3. Vitamin A deficiency caused hyperproliferation of enterocytes, a decrease in the number of goblet cells, decreased alkaline phosphatase activity and decreased expression of 2 brush-border enzymes. 4. Our findings suggest that the absence of vitamin A interferes with the normal growth rate in chickens because it influences functionality of the small intestine by altering proliferation and maturation of cells in the small intestinal mucosa.

Do Physical Forces Contribute to Cryodamage

To achieve the ultimate goal of both cryosurgery and cryopreservation, a thorough understanding of the processes responsible for cell and tissue damage is desired. The general belief is that cells are damaged primarily due to osmotic effects at slow cooling rates and intracellular ice formation at high cooling rates, together termed the “two factor theory.” The present study deals with a third, largely ignored component—mechanical damage. Using pooled bull sperm cells as a model and directional freezing in large volumes, samples were frozen in the presence or absence of glass balls of three different diameters: 70–110, 250–500, and 1,000–1,250 µm, as a means of altering the surface area with which the cells come in contact. Post‐thaw evaluation included motility at 0 h and after 3 h at 37°C, viability, acrosome integrity, and hypoosmotic swelling test. Interactions among glass balls, sperm cells, and ice crystals were observed by directional freezing cryomicroscopy. Intra‐container pressure in relation to volume was also evaluated. The series of studies presented here indicate that the higher the surface area with which the cells come in contact, the greater the damage, possibly because the cells are squeezed between the ice crystals and the surface. We further demonstrate that with a decrease in volume, and thus increase in surface area‐to‐volume ratio, the intra‐container pressure during freezing increases. It is suggested that large volume freezing, given that heat dissipation is solved, will inflict less cryodamage to the cells than the current practice of small volume freezing. Biotechnol. Bioeng. 2009; 104: 719–728

Convergent Effects on Cell Signaling Mechanisms Mediate the Actions of Different Neurobehavioral Teratogens: Alterations in Cholinergic Regulation of Protein Kinase C in Chick and Avian Models

Abstract: Although the actions of heroin on central nervous system (CNS) development are mediated through opioid receptors, the net effects converge on dysfunction of cholinergic systems. We explored the mechanisms underlying neurobehavioral deficits in mouse and avian (chick, Cayuga duck) models. In mice, prenatal heroin exposure (10 mg/kg on gestation days 9‐18) elicited deficits in behaviors related to hippocampal cholinergic innervation, characterized by concomitant pre‐ and postsynaptic hyperactivity, but ending in a reduction of basal levels of protein kinase C (PKC) isoforms βII and γ and their desensitization to cholinergic receptor‐induced activation. PKCα, which is not involved in the behaviors studied, was unaffected. Because mammalian models possess inherent confounding factors from maternal effects, we conducted parallel studies using avian embryos, evaluating hyperstriatal nucleus (intermedial part of the hyperstriatum ventrale, IMHV)‐related, filial imprinting behavior. Heroin injection to the eggs (20 mg/kg) on incubation days 0 and 5 diminished the post‐hatch imprinting ability and reduced PKCg and bII content in the IMHV membrane fraction. Two otherwise unrelated agents that converge on cholinergic systems, chlorpyrifos and nicotine, elicited the same spectrum of effects on PKC isoforms and imprinting but had more robust actions. Pharmacological characterization also excluded direct effects of opioid receptors on the expression of imprinting; instead, it indicated participation of serotonergic innervation. The avian models can provide rapid screening of neuroteratogens, exploration of common mechanisms of behavioral disruption, and the potential design of therapies to reverse neurobehavioral deficits.

Protective effects of iodixanol during bovine sperm cryopreservation

The aim of cryopreservation is to maintain cellular integrity, thereby enabling resumption of proper biological functioning after thawing. Here we propose OptiPrep (60% iodixanol in water) as a protectant during sperm cryopreservation using pooled bull semen as the model. We evaluated OptiPrep concentration effect and its relation to cryopreservation by comparing frozen-thawed and chilled samples. Semen, extended in Andromed with 0 (control), 1.25%, 2.5%, and 5% OptiPrep, was compared after either chilling or freezing in large volume by directional freezing. Sample evaluation included sperm motility upon thawing and after 3h incubation at 37 degrees C for frozen-thawed samples and after 3h and 6h of chilling for chilled samples; viability, acrosomal integrity, and hypoosmotic swelling were also tested for frozen-thawed and chilled samples. Chilled samples with 5% OptiPrep showed inferior viability (P=0.047) and 3h motility (P=0.017) relative to that for chilled samples with 2.5% OptiPrep and inferior viability (P=0.042), acrosomal integrity (P=0.045), and 0h motility (P=0.024) relative to that for chilled samples with 1.25% OptiPrep. The 1.25%, 2.5%, and control samples did not differ. In frozen-thawed samples, 2.5% OptiPrep was superior to all other concentrations for 3h motility (control, P=0.007; 5% OptiPrep, P=0.005; 1.25% OptiPrep, P=0.004) and to 1.25% OptiPrep for acrosomal integrity (P=0.001). In a search for a protection mechanism, we measured glass transition temperature (T(g)) of Andromed and of Andromed with 1.25%, 2.5%, and 5% OptiPrep. Andromed (-58.78 degrees C) and 1.25% OptiPrep (-58.75 degrees C) groups had lower mean T(g) than that of the 2.5% (-57.67 degrees C) and the 5% (-57.10 degrees C) groups. Directional cryomicroscopy revealed that the presence of iodixanol alters ice crystal formation into an intricate net of dendrites. Thus, iodixanol appears to possess cryoprotective properties by helping spermatozoa maintain motility and membrane integrity, possibly through altering ice crystals formation into a more hospitable environment and increasing the glass transition temperature.

The Role of Prolactin in Reproductive Failure Associated with Heat Stress in the Domestic Turkey

Abstract Reproductive failure associated with heat stress is a well-known phenomenon in avian species. Increased prolactin (PRL) levels in response to heat stress have been suggested as a mechanism involved in this reproductive malfunction. To test this hypothesis, laying female turkeys were subjected to 40°C for 12 h during the photo-phase daily or maintained at 24–26°C. Birds in each group received oral treatment with parachlorophenyalanine (PCPA; 50 mg/kg BW/day for 3 days), an inhibitor of serotonin (5-HT) biosynthesis, or immunized against vasoactive intestinal peptide (VIP). Both treatments are known to reduce circulating PRL levels. Nontreated birds were included as controls. In the control group, high ambient temperature terminated egg laying, induced ovarian regression, reduced plasma luteinizing hormone (LH) and ovarian steroids (progesterone, testosterone, estradiol) levels, and increased plasma PRL levels and the incidence of incubation behavior. Pretreatment with PCPA reduced (P < 0.05) heat stress-induced decline in egg production, increase in PRL levels, and expression of incubation behavior. Plasma LH and ovarian steroid levels of heat stressed birds were restored to that of controls by PCPA treatment. As in PCPA-treated birds, VIP immunoneutralization of heat-stressed turkeys reduced (P < 0.05) circulating PRL levels and prevented the expression of incubation behavior. But it did not restore the decline in LH, ovarian steroids, and egg production (P > 0.05). The present findings indicate that the detrimental effect of high temperature on reproductive performance may not be related to the elevated PRL levels in heat-stressed birds but to mechanism(s) that involve 5-HT neurotransmission and the induction of hyperthermia.

The effect of light intensity on growth and development of turkey toms

1. The effect of light intensities from 10 to 700 lux on the performance of 5 to 18 week-old turkey males was studied in 2 trials. 2. Body weight of 18 week-old turkeys, in both experiments, was highest under the lowest light intensity. This coincided with higher weight gain and lower food intake, which resulted in significantly better food conversion efficiency. 3. Light intensity affected heart muscle weight but not weight of breast muscle, abdominal fat or testis as proportions of body weight. 4. The decline in plasma T 3 concentration with age differed from other treatments at the low light intensity, which resulted in a significantly higher T 3 concentration in turkeys exposed to 10 lux at the age of 10 to 15 weeks. 5. It is concluded that light intensity significantly affects food conversion efficiency in turkey males. this is likely to be related to differential investment of energy expenditure for maintenance.