RESULTS: CHICK RESULTS: ZEBRAFISH ABSTRACT In this experiment the effects of ethanol, at concentrations that are physiologically relevant to human alcohol consumption, were tested on vertebrate embryonic heart development. Two model systems that were used are the chick (Gallus gallus) and zebrafish (Danio rerio) embryos. In the chick, exposure to different concentrations of ethanol on heart rate were observed, by pipetting ethanol to unshelled embryos and measuring the heart beats per minute. In the zebrafish model, we observed how ethanol exposure affects heart structure. Freshly fertilized transgenic (cardiac-GFP) zebrafish eggs were placed in Petri dishes containing different concentrations of ethanol. Results indicated that the ethanol exposure in chicks reduced the heart rates significantly, while in zebrafish, physical deformities of the heart and body structure were observed. In both cases, ethanol had adverse effects on embryonic development. INTRODUCTION DISCUSSION/CONCLUSION The Effects of Varying Alcohol Concentrations on Chicken and Zebrafish Embryonic Development Kyla-Gaye Pinnock, Mareneth Banzon, and Arne Christensen BIO 452, Developmental Biology, York College CUNY, Queens, NY 11451, USA The heart is derived from mesoderm and forms in the neck area, moving posterior to the thoracic region as the head elongates. At first, it starts off as a single tube, with the conus arteriosus at the anterior end followed by the atrium, ventricle and ending with the cinus venosus at the posterior. In birds and mammals, the heart takes on a four chambered shape: atrium and ventricle split into left and right, conus arteriosus splits into aorta and pulmonary veins and the cinus venosus develops into the sinoatrial node or pacemaker which regulates heart rate. After 48 hours, the heart of the chick embryo starts to beat. It is at this time, that we performed our experiment with ethanol. Ethanol is a depressant. By binding to nerve receptors, ethanol blocks neurotransmitters from being sent. In the case of the heart, neurons stimulating the sinoatrial node control whether the heart beats faster or slower. Alcohol limits the impulses generated by neurons causing the heart rate to decrease. Our hypothesis is that the embryonic heart rate should slow down after ethanol is given to the embryo and that the effect will increase with the concentration. Zebrafish make good model organisms due to their rapid development and the fact that they can be easily maintained in a ̊C range. While chick embryos can be observed for their heart rate when unshelled, the progress of zebra fish can be seen in a Petri dish so we could compare ethanol exposed embryos to normal embryos. The results are essential in understanding how alcohol affects embryonic development. MATERIALS/METHODS Chick Embryo: A chick egg that was previously incubated for approximately 3 days was cracked open slightly to observe the embryo inside. Increasing concentrations of ethanol – 0.00%, 0.01%,0.05%, or 0.09%, were added to the embryo and the heart rate was subsequently recorded. Zebrafish Embryo: Transgenic zebrafish Tg(cmlc2:egfp) embryos with a heart-specific promoter driving cardiac GFP expression were provided by Dr. Nathalia Glickman Holtzman from Queens College / CUNY. Viable eggs were selected from stock eggs, and were placed in egg water containing increasing concentrations of ethanol; 0.0, 0.5, or 1.0 %. Approximately 40 eggs were used for each treatment, and observations were made five days after the introduction of alcohol into the systems of the zebrafish. Alcohol Negatively affects both chick and zebrafish embryos High concentrations of ethanol showed reduced heart rate in chick embryos (Figure 1, 0.09% ethanol 10 minutes after treatement). Ethanol induced developmental defects in zebrafish including fattened tails, a decrease in responsiveness, swelling of the pericardial sac, and the elongation of the heart atrium and ventricle Concrete conclusions cannot be drawn from one experiment but it was observed that alcohol causes developmental defects in both chick and zebrafish embryos. REFERENCES Gilbert, S. F. (2010). Developmental Biology (9th ed.). N.p.: Sinauer Associates, Inc. Tyler, M. S. (2010). Developmental Biology: A Guide For Experimental Study (3rdrd ed.). Saunderland, MA: Sinauer Associates, Inc. Dlugos, C.A and Rabin, R. A. (2010). R.A Structural and Functional Effects of Developmental Exposure to Ethanol on the Zebrafish Heart. Alcohol Clin Exp Res. 34, Effects of ethanol on heart rate (BPM) Figure 1: Chick embryos were treated with different concentrations of ethanol (EtOH) – 0.00, 0.01,0.05, or 0.09 %. Heart rate was determined before treatment, immediately after treatment, and 10 minutes after treatment. Heart rate was determined five times and then averaged (last table column). Figure 2: Pictures of the zebrafish heart expressing Green Florescent Protein. A: The left panel shows a regular two chambered zebrafish heart. B: The right panel shows the deformed heart, and enlarged pericardial sac, of a zebrafish exposed to ethanol. C: The left panel shows normal zebrafish heart structure. D: The right panel shows the deformed heart of a zebrafish exposed to ethanol. The atrium and the ventricle are somewhat indistinguishable.