1. Adenosine and Dopamine Receptor Interactions in the Striatum and Caffeine-Induced Behavioral Activation Vickram Ramkumar, Ph.D. and Linda Toth, DVM, Ph.D. Department of Pharmacology Southern Illinois University School of Medicine Springfield, IL

2. ARROS Antioxidant Enzymes Nuclear Factor  B Adenosine Receptor – Reactive Oxygen Species Interaction (ROS) Are Adenosine Receptors Markers of Oxidative Stress?

3.  Chemotherapeutic drugs which increase ROS generation increase adenosine A 1 receptor by activating nuclear factor  B (Nie et al., 1998)  Nerve growth factor induces the expression of the adenosine A 2A receptor by activating nuclear factor  B (Nie et al., 1999)  NF-κB induces the expression of the A 2A AR while A 2A AR suppresses NF-kB activation (Sands et al., 2004; Murphee et al., 2008)  Deletion of the p50 subunit of NF-  B decreased A 1 AR and D 2 R expression and increased A 2A AR expression (Jhaveri et al., 2007; Xie et al., 2008)  NF-κB dependent induction of dopamine D 2 receptor (Fiorentini et al., 2002) Previous Observations Conclusion: NF-kB plays an integral role in the regulation of adenosine and dopamine signaling

4. Cisplatin Oxidative Stress Induces Cochlear A 1 AR Ford et al., 1997 Hear Res. 111, 143-152

5. OXIDATIVE STRESS INDUCES A I ADENOSINE RECEPTORS Nie et al., 1998 Mol. Pharmacol. 53, 663-669

6. Cisplatin Induces NF-  B Activation Nie et al., Mol. Pharmacol. 53, 663-669

7. Nerve Growth Factor suppresses A 2A AR expression through activation of NF-κB Nie et al., 1999; Mol. Pharmacol. 56, 947-954

8. What is NF-kappa B? (A 1 AR,. A 2A AR) ROS (Modified from)

9. Does Disruption of NF-  B Function Alter Expression of A 1 and A 2A AR? Hypothesis: Disruption of NF-κB would lead to lower A 1 AR and higher A 2A AR expression. (B6129PF2/J)(B6129P-Nfkb1 (tmbl)

10. Decreased Expression of A 1 AR in the Mouse Striatum Xie et al., 2007 Life Sci. 1031-1041

11. Increased Striatal Expression of A 2A AR in p50 -/- Mouse Xie et al., 2007 Life Sci. 1031-1041

12. Decreased Striatal Expression of D 2 Dopamine Receptor in p50 -/- Mouse Xie et al., 2007 Life Sci. 1031-1041

13. Regulation of G Proteins in the p50 -/- Mouse Striatum Xie et al., 2007 Life Sci. 1031-1041

14. SUMMARY  Decreased expression of the A 1 AR  Increased expression of the A 2A AR  Decreased expression of D 2 R  Increased G olf/s proteins & decreased G i

15. Striatal Adenosine/Dopamine Receptors Distribution

16. Purported AR/DR Signaling Cascade in Striatum G olf/s GiGi A 1 AR A 2A AR D2RD2R D1RD1R Cyclic AMP Protein Kinase A CREB IEGs GABAergic Neuron Activity

17. Do Alterations in Striatal Adenosine and Dopamine Receptors in p50 - /- Mouse Striatum Confer Behavioral Differences?

18. (300 mg/L) F2: 101 mg/kg/24 h; p50-/- 107 mg/kg/24 h (20 g/L) (Intraperitoneal transmitter and DSI receiver under each cage)

19. Increased Sensitivity of p50 -/- Mice to the Locomotor Stimulatory Action of A 2A AR Antagonist (10 mg/kg, i.p.) (5 mg/kg, i.p.)

20. D 2 R Antagonist Reversed the Stimulatory Action of A 2A AR Antagonist Raclopride (0.5 mg/kg, i.p.) SCH 58621 (10 mg/kg, i.p) DPCPX (5 mg/kg, i.p)

21. A. B. C.D. 0 2 4 6 8 10 12 14 16 14 Hours after oral administration Serum Caffeine (ug/ml) 0 2 4 6 8 10 14 Hours after oral administration Caffeine in Brain (ug/g) 0 2 4 6 8 10 12 14 16 051015202530 Minutes after injection Caffeine in brain (ug/g) F2 KO F2 (n = 4) KO (n = 3) ● ┼ KO, n = 6 F2, n = 3-6 No Change in Caffeine Levels in Serum and Brain between F2 and p50 -/- Mice

23. Summary  p50 -/- mice do not show any differences in basal locomotor activity.  p50 -/- mice are more sensitive to the locomotor stimulatory effect of caffeine and A 2A AR antagonist but not A 1 AR antagonist.  p50 -/- mice show reduced dopamine turnover but no change in dopamine transporter or altered caffeine pharmacokinetics.

24. AR/DR Signaling in p50 -/- Mouse Striatum

25. Adenosine/Dopamine Neurotransmission in the Striatum

26. Receptors G protein/ second messengers Kinases Changes in p50-/- Striatum Behavior

27. Acknowledgements Linda Toth, DVM, Ph.D. Xiaobin Xie, Ph.D. Krishna Jhaveri, Ph.D. Lydia Abrogast, Ph.D. Zhongzhen Nie, M.D., Ph.D – Univ. Georgia, Augusta, GA Gary Stiles, M.D. – Duke University, Durham, NC Honzhou Ren, Ph.D. – Duke University, Durham, NC NIH Grants NIHLB, NIDCD, SIU CRC Funds