Description) Neonatal Intensive Care exposes 260,000 preterm neonates, born in the U.S. each year, to prolonged pain, stress and discomfort. Follow-up studies of ex-premature neonates show major behavioral disorders, altered pain thresholds, attention deficit disorder, severe anxiety, high somatization scores, and learning deficits. The investigators hypothesize that prolonged neonatal pain and stress may lead to changes in neuronal and neurotransmitter development which result in these behavioral disorders. The investigators propose to develop models of clinical pain using newborn rat pups, and record their vocalizations and pain-related behaviors at different postnatal ages in response to episodic or repetitive pain. Neuronal activation from painful stimuli at different ages will be mapped by the expression of c-Fos as a trans-synaptic marker using in situ hybridization (ISH) and immunocytochemistry (ICC). Co-expression of c-Fos and genes associated with neuronal plasticity (GAP43 and HSP-70) will help to probe the effects of painful versus tactile stimulation on development in the sensory cortex, hippocampus, thalamus and hypothalamus. RNAse protection assays will be used for initial estimation of gene expression, followed by ISH and ICC to examine cellular detail and co-expression. Differential effects of inflammatory or thermal pain, effects of increasing pain intensity and repetitive pain will be tested in subsequent experiments. In later childhood, ex-preterm neonates manifest altered pain-related behaviors, severe anxiety, and learning deficits. Thus, rat pups subjected to repetitive pain will be reared to adulthood and tested by the defensive withdrawal test, air-puff startle response, Morris swim maze test, the social discrimination test, and the alcohol preference test. Hypothalamic-pituitary-adrenal axis responses and pain thresholds will be measured (hot plate test) at baseline and after restraint stress to examine pain behaviors and stress-induced analgesia. Adult behavioral responses will be correlated with the expression of c-Fos, GAP-43, and the density of u-s- and K-opioid receptors in the areas outlined above. Robust correlations between behavior patterns and neuronal gene expression will generate mechanistic hypotheses to investigate the neurobiologic basis for behavior following neonatal pain/stress. These studies may have important implications for the behavior and the health-care problems of large numbers of ex-premature infants approaching adolescence and adulthood in the next decade. The Stress Neurobiology Laboratory will train Dr. Anand in the techniques of molecular neurobiology and help in performing these experiments. Active investigations include: central regulation of the HPA axis, effects of early experience on HPA responsiveness, hypothalamic control of immune-endocrine interactions and neuroendocrine function. These studies use a variety of behavioral, neuroendocrine, and molecular techniques under the direction of scientists with established track-records in basic research and extra-mural funding.
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