Neuroscience Refresher Course The Brain and the Immune System François M. Abboud, MD University of Iowa Carver College of Medicine “IT’S ALL IN YOUR HEAD” Experimental Biology Meeting Boston Convention Center Saturday, March 28, 2015

Part I: The Power of the Autonomic Nervous System Part II: Neuro-Immune Synapse

Neuronal Signals Are Powerful Regulators of Cardiovascular Disease Nobel Prize 1938 Corneille Heymans Discovery of the role played by sinus and aortic mechanisms in the regulation of respiration Nobel Prize 1970 Katz, von Euler, Axelrod Discovery of humoral transmitters in nerve terminals and the mechanisms for their storage, release and inactivation

Movie clips of people fainting in different situations

Fig. 1. From Wallin and Sundlof. J Aut Nerv Sys 6(3): , Available at sciencedirect.com/science/article/pii/ sciencedirect.com/science/article/pii/

Cardiac Vagal Afferent Reflex Neurocardiogenic Syncope Sympathetic Efferents Medullary Cardiovascular Centers Vagal Afferents Sympathoinhibition Bradycardia

Autonomic Regulation Old Concept: Autonomic regulation is essential for acute circulatory adjustment New Concept: Autonomic dysregulation induces pathological processes that determine cardiovascular mortality Plasma NE levels positively correlate to increased mortality in heart failure Fig. 3. From Cohn et al. New Engl J Med 311(13): , Available at nejm.org/doi/pdf/ / NEJM nejm.org/doi/pdf/ / NEJM

Direct Recordings of Sympathetic Nerve Activity in Patients with Heart Failure (Dramatic Increases Seen with Severe Failure) Fig. 2. From Ferguson et al. J Am Coll Cardiol 16: , Freely available at content.onlinejacc.org/article. aspx?articleID= content.onlinejacc.org/article. aspx?articleID= Figs. 1, Left, & 4. From Leimbach et al. Circulation 73: , Freely available at circ.ahajournals.org/cont ent/73/5/913.full.pdf+html circ.ahajournals.org/cont ent/73/5/913.full.pdf+html

Baro-stimulation Produces Sustained Hypotension and Bradycardia in Conscious Dogs Fig. 1. From Lohmeier et al. Hypertension 43(2): , Freely available at hyper.ahajournals.org/content/ 43/2/306.full hyper.ahajournals.org/content/ 43/2/306.full

Improved Survival in Heart Failure Figs. 7 & 5. From Zucker et al. Hypertension 50: , Freely available at hyper.ahajournals.org/ content/50/5/904.full hyper.ahajournals.org/ content/50/5/904.full Figs. 4 & 5C. From Li et al. Circulation 109(1): , Freely available at circ.ahajournals.org/content /109/1/120/F5.expansion.ht ml circ.ahajournals.org/content /109/1/120/F5.expansion.ht ml

Autonomic Dysregulation Increases Cardiovascular Mortality Poor outcomes in Cardiovascular Disease Decreased Parasympathetic Increased Sympathetic Can it be Reversed in Humans?

Vagal Stimulation in Heart Failure Current Clinical Trials Nerve Stimulation Carotid Sinus Nerve Stimulation in Hypertension Baroreflex – Sustained Hypotension/Bradycardia Control1234 Volts

Part I: The Power of the Autonomic Nervous System Part II: Neuro-Immune Synapse (Fatal Conspiracy in Cardiovascular Disease)

The Neuro-Immune Triangle in Cardiovascular Disease Autonomic Dysregulation Increases Mortality Autonomic System is a Powerful Regulator of the Immune System Inflammatory/Immune System Causes CV Damage DeathLife A BC

Immune System Contributes Extensively to Cardiovascular Damag e Atherosclerosis –Plaques composed of foam cells, T-cells, and dendritic cells Heart Failure –Cytokine “storms” Atrial Fibrillation –Fibrosis  Remodeling Stroke –Macrophage extravasation peroxiredoxin release Hypertension –Innate and Adaptive immune responses

Hence ……. NO THYMUS NO delayed HTN Immune System & Hypertension Control + DOCA/SALT HTN Experimental + DOCA/SALT NO HTN Svendsen UG. Acta Pathol Microbiol Scand A 84(6): , Available at onlinelibrary.wiley.com/doi/ /j tb00150.x/pdfonlinelibrary.wiley.com/doi/ /j tb00150.x/pdf

Role of Immune System in Hypertension Fig. 4. From Ba et al. J Immunol 128(3): , Freely available at jimmunol.org/content/128/3/1211.full.pdf+html Thymic Transplant in SHR

Ang II Hypertension Mediated by T-cells Figs. 5 & 4B (top 2 and bottom lines only). From Guzik et al. J Exp Med 204(10): , Freely available at jem.rupress.org/content/204/10/24 49.full

IL-6 Knockout Prevents Ang-II Hypertension Figs. 1 (top 2 lines) & 3. From Brands et al. Hypertension 56: , Freely available at hyper.ahajournals.org/content/56/5/ 879.full. hyper.ahajournals.org/content/56/5/ 879.full Arterial Pressure Afferent Renal Arteriole

Ang II Increases Renal JAK2 and STAT-3 Phosphorylation in WT But Not in IL-6 KO JAK2STAT-3 Fig. 4. From Brands et al. Hypertension 56: , Freely available at hyper.ahajournals.org/content/56/5/ 879.full. hyper.ahajournals.org/content/56/5/ 879.full

Figs. 1 & 3. From Ganta et al. Am J Physiol Heart Circ Physiol 289: H , Reprinted with permission. Freely available at ajpheart.physiology.org/content/289/4/H1683ajpheart.physiology.org/content/289/4/H1683

SNS induces release of Hematopoietic endothelial stem and progenitor cells from BM to the spleen where SCF and accelerates monocyte production Myocardial Infarction Accelerates Atherosclerosis Dutta et al. Nature Med 18: 902–910, Available at nature.com/nature/journal/vaop/ncurrent/abs/nature11260.htmlnature.com/nature/journal/vaop/ncurrent/abs/nature11260.html

Migration of HSPC from the Bone Marrow by Sympathetic Activity (Post-MI) Fig. 3. From Swirski FK and Nahrendorf M. Science 339: , Available at sciencemag.org/content/339/6116/161.full sciencemag.org/content/339/6116/161.full

Circadian Oscillation in Sympathetic Nerve Activity Govern Recruitment of HSPCs to Bone Marrow and Leukocytes to Muscle Scheiermann et al. Immunity 37: , Freely available at sciencedirect.com/science/article/pii/S sciencedirect.com/science/article/pii/S

Fig. 1. From O’Mahony et al. Am J Physiol Regul Integr Comp Physiol 297(4): R1118-R1126, Reprinted with permission. Freely available at ajpregu.physiology.org/content/297/4/R1118 ajpregu.physiology.org/content/297/4/R1118 Fig. 1A. From Karimi et al. Brain Behav Immun 24: , Available at sciencedirect.com/sci ence/article/pii/S sciencedirect.com/sci ence/article/pii/S Tonic Suppressive Vagal Influence on the Immune System

Vagal Nerve Stimulation Attenuates the LPS-Induced TNF Response and Endotoxin Shock Fig. 1. From Tracey KJ. Nature 420: , Available at nature.com/nature/journal/ v420/n6917/full/nature html nature.com/nature/journal/ v420/n6917/full/nature html Fig. 3. From Borovikova et al. Nature 405: , Available at nature.com/nature/journal/ v405/n6785/full/405458a0. html nature.com/nature/journal/ v405/n6785/full/405458a0. html “The mind has great influence over the body, and maladies often have their origin there.” Moliére

Fig. 4, B-F. From Li et al. Hypertension 57: , Freely available at hyper.ahajournals.org/content/57/2/298.full hyper.ahajournals.org/content/57/2/298.full Proinflammatory State in 2K1C Hypertension Is Enhanced in  -7 Nicotinic Receptors KO Mice IL-6 IL-1  TNF-  LVW/BWGlomerular Sclerosis

 -7 Nicotinic Receptors KO Mice Proinflammatory State in 2K1C Hypertension Fig. 5, A & B, right. From Li et al. Hypertension 57: , Freely available at hyper.ahajournals.org/content/57/2/298.full hyper.ahajournals.org/content/57/2/298.full

Is there an intrinsic abnormality in the response of innate immune cells from WKY vs. SHR to known immunological Toll-like Receptor ligands? Do angiotensin and the cholinergic neurotransmitter nicotine alter the immunological response of immune cells from WKY vs. SHR?

Hypothesis: Abnormal Autonomic Immunomodulation TLR2, TLR3, TLR4, TLR5, TLR7/8, & TLR9 Adapted from Fig. 1. From Tracey K. Nature Rev Immunol 9: Available at nature.com/nri/journal/v9/n6/full/nri2566.htmlnature.com/nri/journal/v9/n6/full/nri2566.html

TLR Ligands WITH Nicotine OR Ang II TLR Ligands Alone Cytokine Release from Rat Splenocytes WKY = Normotensive Rat SHR = Pre-hypertensive Rats were 3-6 weeks old Supernatants were collected for measurements of TNF-alpha, IL-6, IL-10 by ELISA. A dose response to each TLR ligand was obtained first. Methods & Protocols

Cytokine Responses to TLR Activation Fig. 1. From Harwani et al. Circ Res 111: , Freely available at circres.ahajournals.org/content/ 111/9/1190.full

IL-6 Release Is Enhanced in SHR Fig. 2. From Harwani et al. Circ Res 111: , Freely available at circres.ahajournals.org/content/ 111/9/1190.full circres.ahajournals.org/content/ 111/9/1190.full

Methods and Protocols In Vivo Experiments Osmotic pumps containing nicotine or saline were implanted. Infusions were conducted over 24 hours. At 20 hours, animals received an intraperitoneal injection of either sterile saline or Clo 97 (a TLR7/8 ligand). At 24 hours, sera was collected and assayed for cytokines.

IL-6 and IL1  Levels Are Increased in SHR and Suppressed in WKY by Nicotine Fig. 3, E & F. From Harwani et al. Circ Res 111: , Freely available at circres.ahajournals.org/content/1 11/9/1190.full circres.ahajournals.org/content/1 11/9/1190.full In Vivo Serum Levels

Activated Macrophages Increase With Nicotine in SHR Fig. 4. From Harwani et al. Circ Res 111: , Freely available at circres.ahajournals.org/con tent/111/9/1190.full circres.ahajournals.org/con tent/111/9/1190.full

Summary 1.The autonomic nervous system exerts a powerful modulatory influence on the immune system with pro-inflammatory morbid cardiovascular consequences. 2.Vagus nerve activity provides a protective anti-inflammatory effect mediated by  7-nicotinic cholinergic receptors. 3.In a genetic model of hypertension (SHR), the anti-inflammatory effect of nicotine on innate immune cell is reversed to a pro-inflammatory response prior to the onset of hypertension. 4.An excessively large population of CD161+ splenocytes is present in SHRs in the neonatal state and increases with age as hypertension progresses

Conclusion The innate immune system in genetic hypertension is abnormally regulated by the autonomic nervous system to trigger pro- inflammatory responses to endogenous antigens. These induce pathologic renal and vascular changes that initiate and sustain the hypertensive state.

Acknowledgements Mark Chapleau Sal Harwani Madhu Singh Yongjun Lu Mike Czicha Carol Whiteis Immunologists: Zuhair Ballas, Kevin Legge, Nick Zavazava and Fayyaz Sutterwalla Consultants: David Harrison and Howard Jacob Core Facilities: Cell Sorting and Gene Transfer Vector

François M. Abboud, MD University of Iowa Carver College of Medicine François M. Abboud, MD Associate Vice President for Research Edith King Pearson Chair of Cardiovascular Research Chair Emeritus, Dept. of Internal Medicine ( ) Founding Director, Francois M. Abboud Cardiovascular Research Center ( ) Professor of Internal Medicine - Cardiovascular Medicine Professor of Molecular Physiology and Biophysics Learn more at ppointment=Internal%20Medicine&id=abboudf ppointment=Internal%20Medicine&id=abboudf To receive a complete set of unaltered slides, please contact the APS Education Office