Neuroscience Scholars Summer Program Project

1. Neuroscience Scholars Summer Program Project Kirsten R. Gambrell Summer 2005 Mentors: Dr. Elise Gomez-Sanchez Dr. Celso Gomez-Sanchez

2. The Gomez-Sanchez Lab –General Questions about Hypertension How do steroids of the adrenal cortex induce hypertension? Particularly, how does the mineralocorticoid aldosterone alter the central control of the blood pressure? How is the synthesis of steroids of the adrenal cortex (aldosterone and the glucocorticoids, cortisol and corticosterone) regulated?

3. Mineralocorticoid Receptor (MR) MR transcription factor Promotes transcription of different genes depending on the cell type and binding responsible for sodium and water transport in the kidney & other transport epithelia The MR is activated by binding to an agonist

6. Brain MR and its link to hypertension The affinity of the MR for its primary ligands is tissue dependent 11b-hydroxysteroid dehydrogenase provides extrinsic specificity for aldo in the kidney. Cortisol & corticosterone hippocampus Aldosterone Hippocampus Paraventricular n (hypothal) Amygdala Cortex cerebellum The mechanism for ligand preference in the brain is not known The circumventricular organs & amygdala are involved with blood pressure and fluid and electrolyte homeostasis.

8. Specific Questions for NSSP Do oxytocin and vasopressin have a direct relationship with MR? Do oxytocin and vasopressin receptors co-localize with those of MR? Does RACK1 protein interact with the MR and give it ligand specificity?

9. MR Slides

10. Oxytocin and Vasopressin Synthesized in the PVN and SON of the hypothalamus and released into the general circulation from the pituitary Nine-aa, peptide neurotransmitters similar in aa sequence but different in function Oxytocin: uterine contraction, milk let-down, and blood pressure control, counteracting the affects of vasopressin Vasopressin: vasoconstriction and renal fluid absorption

12. Protocol Neuroanatomy Selective expression for vasopressin, oxytocin and MR Rat brain – hypothalamus, specifically PVN and SON IHC Bathing brain sections in the primary Ab (MR, oxytocin, vasopressin) then secondary Ab Staining both with DAB and immuno-fluorescence

13. Vasopressin Stained Slides

14. Oxytocin Double Stained Slides

15. Results Based on double stained slides, both vasopressin and oxytocin are expressed in the same cell. Sharp 3D images of the slides using either a confocal microscope or de-convolution software with a Z-motor will confirm these results. This suggests that vasopressin and oxytocin may interact directly with MR

16. Conclusion What does this mean? In the normal animal model, we found that there is a relationship between vasopressin and MR, as well as oxytocin and MR. What’s next? These relationships will be studied in animal models, both genetic and pharmacologic, of human hypertension, including salt-sensitive hypertension, that are responsive to MR antagonists.

17. RACK1 RACK1 is a scaffolding protein that co-precipitates in a yeast-2-hybrid assay with the MR. The hypothesis is that RACK1 alters the configuration of MR, thus increasing its specificity for aldosterone

18. Special Thanks to … Dr. Elise Gomez-Sanchez Dr. Celso Gomez-Sanchez Angela F de Rodriguez Miriam Gomez-Sanchez Dr. Ian Paul Detra Scott Ann Dautenhahn Gomez-Sanchez Lab Neuroscience Summer Scholars