Developmental expression of Kv1 voltage-gated potassium channels in the avian hypothalamus.

Specialized hypothalamic neurons integrate the homeostatic balance between food intake and energy expenditure, processes that may become dysregulated during the development of diabetes, obesity, and other metabolic disorders. Shaker family voltage-gated potassium channels (Kv1) contribute to the maintenance of resting membrane potential, action potential characteristics, and neurotransmitter release in many populations of neurons, although hypothalamic Kv1 channel expression has been largely unexplored. Whole-cell patch clamp recordings from avian hypothalamic brain slices demonstrate a developmental shift in the electrophysiological properties of avian arcuate nucleus neurons, identifying an increase in outward ionic current that corresponds with action potential maturation. Additionally, RT-PCR experiments identified the early expression of Kv1.2, Kv1.3, and Kv1.5 mRNA in the embryonic avian hypothalamus, suggesting that these channels may underlie the electrophysiological changes observed in these neurons. Real-time quantitative PCR analysis on intact microdissections of embryonic hypothalamic tissue revealed a concomitant increase in Kv1.2 and Kv1.5 gene expression at key electrophysiological time points during development. This study is the first to demonstrate hypothalamic mRNA expression of Kv1 channels in developing avian embryos and may suggest a role for voltage-gated ion channel regulation in the physiological patterning of embryonic hypothalamic circuits governing energy homeostasis.

The effect of cAMP signaling on the longitudinal extension of spinal sensory neurons in the chicken embryo.

Developing sensory axons grow into the spinal cord in a three-step process: the axons extend toward and into the cord, then branch rostrally and caudally to establish a longitudinal pathway, and finally grow into the grey matter. This study investigated regulation by cAMP of the longitudinal extension of this pathway within the spinal cord. The cAMP pathway was pharmacologically altered in chicken embryos to determine its effects on the establishment of the longitudinal extension of the dorsal funiculus. A forskolin-induced increase in cAMP in ovo inhibited longitudinal growth by sensory afferents. Furthermore, blocking cAMP activation of protein kinase A (PKA) in ovo with H-89 substantially increased longitudinal extension. These results demonstrate a specific role for the cAMP/PKA pathway in the initial longitudinal spinal afferent growth in the chicken embryo.

An in ovo chicken model to study the systemic and localized teratogenic effects of valproic acid.

BACKGROUND: The antiepileptic valproic acid (VPA) is a teratogen whose embryopathic mechanism(s) remain uncertain. Elucidating potential cellular and molecular effects of VPA is complicated by systemic application paradigms. We developed an in ovo model to reproduce the teratogenic effects of VPA and a localized VPA application procedure to determine whether VPA can selectively effect abnormal development in one region of the embryo. METHODS: VPA was applied topically to chicken embryos in ovo at different embryonic stages. Embryos were later evaluated for gross and skeletal anomalies. Pax-2 and Pax-6 protein expression in the developing eye was also evaluated because VPA-induced eye anomalies are similar to those seen by the disruption of Pax-2 and Pax-6. For localized application, a thin sheet of the synthetic polymer Elvax was impregnated with VPA. A small piece of the VPA-impregnated polymer was applied directly to the presumptive wing bud region in Stage 10-17 embryos. Embryos were examined for gross and skeletal anomalies. Sham controls were employed for all experiments. RESULTS: Chicken embryos exposed to VPA in ovo demonstrated increased mortality, growth delay and anomalies similar to ones previously seen in humans: neural tube, cardiovascular, craniofacial, limb and skeletal. Pax-2 and Pax-6 protein expression was qualitatively diminished in the eye. Localized wing bud VPA exposure caused structural abnormalities in the developing wing in the absence of other anomalies in the embryos. These wing defects were similar to those observed after topical whole-embryo VPA application. CONCLUSIONS: These results indicate that at least one mechanism for the teratogenicity of VPA involves a direct effect on developing tissue. The nature of the abnormalities observed implies that this effect may be mediated by disruption of genes that regulate pattern formation.