The Physiology of Oxytocin and Its Role in Social Behavior and Health

Philip McCabe

Department of Psychology

Health Psychology Program;

Neuroscience Program

University of Miami

Coral Gables, FL

Oxytocin

Oxtocin (OT) means “Quick Birth” OT found in placental mammals, some

marsupial mammals, and the ratfish Related peptides found in most other

species (e.g., isotocin)

Oxytocin: Chemical Properties 9 amino acid peptide Molecular weight is 1007 Differs from vasopressin (AVP) by

amino acids at positions 3 and 8

Oxytocin: Biosynthesis Human OT gene mapped to chromosome

20p13 Gene transcribes inactive precursor, OT-

Neurophysin I, which is hydrolyzed by enzymes into smaller fragments, including OT and Neurophysin

Gimpl & Fahrenholz, 2001

Hypothalamic Source of OT OT synthesized in hypothalamic

magnocellular neurons in the paraventricular nucleus (PVN) and supraoptic nucleus (SON)

OT also found in some parvocellular neurons in PVN

Hypothalamic OT Cell Bodies

3V

PVN

SON

Optic Chiasm

Ludwig & Leng, Nature Reviews Neuroscience, 2006

Neurohypophyseal Mechanism

OT: Beyond Reproduction Classic OT functions related to

parturition and milk ejection in females OT found in equivalent concentrations

in the posterior pituitary and plasma of both sexes

Suggests OT has functions beyond female reproductive functions

Plasma Titers of OT Human 0.13pg/ml-414pg/ml Monkey 5.0pg/ml-275pg/ml Cow 1.0pg/ml-10.0pg/ml Rabbit 8.0pg/ml-3000pg/ml Rat 1.0pg/ml-700pg/ml Guinea Pig 1.0pg/ml-25pg/ml Vole 250pg/ml-500pg/ml Mouse 1.0pg/ml-300pg/ml

OT Release and Metabolism Evidence that OT is released in a pulsatile

fashion Turnover rate of OT is fast

Half life in plasma reported between 2-12 minutes (most estimates between 2-7 minutes)

Half life in brain is longer (approx. 30 minutes) OT is metabolized by an enzyme, oxytocinase At physiological concentrations, OT does not

appear to cross the blood-brain-barrier

OT Tissue Expression OT is also expressed in a variety of

tissues, including: Ovaries/Corpus Luteum, Uterus, Placenta,

Prostate Gland, Testes/Leydig Cells, Thymus, Adrenal Medulla, Heart, Aorta, Cancer Tumors, Brain

OT may have local autocrine/paracrine actions

Oxytocin Receptor (OTR) OTR is a 389 amino acid polypeptide 7 transmembrane domains G-protein coupled receptor OTR gene is present as single copy

mapped to chromosome 3 in humans Forms a subfamily of structurally related

receptors with 3 vasopressin receptor subtypes (V1a, V1b, and V2)

Gimpl & Fahrenholz, 2001

Oxytocin Receptor

Gimpl & Fahrenholz, 2001

OTR Selectivity OTR is relatively non-selective OTR has only a 10-fold higher affinity

for OT than AVP AVP acts as partial agonist for OTR Need 100-fold greater concentration of

AVP than OT to get a comparable response

Gimpl & Fahrenholz, 2001

OTR Signaling Pathways OTRs are functionally coupled to Gq/11α class GTP

binding proteins that stimulate phospholipase C-β isoforms

Leads to production of inositol triphosphate, which triggers Ca++ release from intracellular stores, and diacylglycerol

This stimulates protein kinase C, which phosphorylates target proteins and activates the ERK1/2 pathway

Intracellular Ca++ leads to production of nitric oxide and cGMP

OTR Signaling Pathways

Devost, Wrzal & Zingg, 2008

OTR Regulation OTRs go through dramatic tissue-specific up

and down regulation (as much as 10-100 fold)

Regulation occurs at the transcriptional, translational and protein levels

Down regulation of OTR can occur in seconds/minutes as receptor uncouples from the G-protein and undergoes endocytosis, internalization and sequestration

Devost, Wrzal & Zingg, 2008

OTR Regulation OTRs require two elements for high

affinity binding: divalent cations (Mg++ or Mn++) and cholesterol (membrane stabilization)

Estrogen up regulates the expression of OTRs, whereas progesterone inhibits the action of OTRs

OTRs: Acute-Phase Response Elements?

Promoter region of OTR gene contains IL-6 response elements and acute- phase response elements

Suggests that the acute induction of OTR expression could be similar to induction of acute-phase response genes induced by infection or inflammation

OTR Tissue Expression OTRs are expressed in a variety of

tissues, including: Specific brain regions, uterus, mammary

glands, pituitary gland, prostate gland, heart, blood vessels, kidney, pancreas, adrenal gland, cancerous tumors (e.g., breast, uterus, brain, lung), lymphocytes, macrophages and adipocytes

No OTRs observed in liver

Role of OT in Social Behavior and Brain Activity

Beginning in 1970’s, OT implicated in coordination of behaviors in mothers necessary for survival of offspring (i.e., maternal behavior)

Accumulating evidence that OT acts in the brain to modulate constellation of behaviors associated with sociality (social cognition & affiliative behavior in both sexes; see Ross & Young, 2009 for review)

Maternal Care in Rodents OT injected intracerebroventricularly (i.c.v.) in virgin

rats elicited maternal behavior toward pups within 2 hours (facilitated by estrogen priming)

i.c.v. administration of OT antagonists (OTAs) blocked maternal behavior in rats who just gave birth

OT plays more important role in the initiation of maternal behavior than the maintenance of these behaviors

OT knockout mice (OTKO) and OTR knockouts (OTRKO) support these pharmacological findings

Ovine Maternal Bonding Sheep form strong selective mother-lamb social

bonds, and mothers will reject lambs who are not their offspring

This effect seems to be dependent on olfactory memory

i.c.v. OT induces maternal behavior in less than one minute in estrogen-primed nonpregnant ewes, and facilitates olfactory memory by modulating norepinephrine and synaptic plasticity in the olfactory bulb

Alloparental Behavior in Voles Unlike most rodents and mammals, prairie

voles are socially monogamous, and form selective preference for one mate (i.e., pair bond)

Prairie voles also display biparental care, and will “baby sit” vole pups not their own

There is evidence that OTR density in the nucleus accumbens (NAcc) of the brain is related to this alloparental care

OTR Density in NAcc and Alloparental Behavior (Olazabal & Young, 2006)

Social Bonding in Adult Voles i.c.v. infusion of OT during 6 hr cohabitation

with male induces partner preference in unmated female voles

OTA administration in mated females blocks pair bonding

i.c.v. AVP in males induces partner preference

AVP antagonist blocks pair bonding in males Role of OT in male pair bonding not clear

Ross & Young, 2009

OT and AVP in Pair Bonding

Young, Wang & Insel, 1998

Pair Bonding and NAcc OTRs Monogamous prairie voles have greater density of

OTRs in NAcc than non-monogamous species of voles

OTA injected into NAcc in female prairie voles prevented the formation of a partner preference

Over-expression of OTRs in NAcc enhanced pair bonds in prairie voles, but not other species of voles

Therefore, increased OTR in NAcc alone is not sufficient to produce species differences in pair bonding

Ross & Young, 2009

Pair Bonding and NAcc OTRs

Insel & Young, 2001

Why Is NAcc OT Important? NAcc receives dopamine input from midbrain ventral

tegmental area, and is part of mesolimbic dopamine reward/reinforcement pathway

It has been shown in female prairie voles that dopamine and OT systems interact in NAcc to promote pair bonding (i.e., reinforcement of the mating experience)

It has been suggested that ability of female to form attachment with male partner evolutionarily arose from a modification of the neural machinery involved in regulating maternal behavior

Ross & Young, 2009

Social Recognition in Rodents It has been hypothesized that pair bond

formation is due to an association between the rewarding mating experience and the olfactory signature of the partner (i.e., social recognition)

i.c.v. OT increases the amount of time a male rat remembers a conspecific

Brain Mechanisms in Social Recognition

Brain regions implicated in rodent social memory are: Ventral Hippocampus, Septal Nuclei, Medial

Preoptic Area and Olfactory Bulb Although OT injected into these regions

enhances memory, OTA administration does not block memory performance, and therefore the brain mechanisms for social recognition are not clear

OTKO Mice and Social Memory

OTKO mice show deficits in social memory, but not general memory deficits (e.g., habituation to non-social odors)

A single i.c.v. injection of OT before initial exposure completely rescues the deficit in social recognition

Effect seems to involve the medial amygdala OT may act to enhance the saliency of social

stimuli and to encode social memories, which facilitates social relationships

Ross & Young, 2009

Neural Circuitry of OT System Cell bodies of OT neurons are almost

exclusively confined to the hypothalamic PVN and SON

OT fibers are located throughout the CNS: Dorsal Medial Hypothalamus, Several Thalamic

Nuclei, NAcc, Hippocampus, Entorhinal Cortex, Septal Nuclei, Amygdala, Olfactory Bulbs, Periaqueductal Grey, Substantial Nigra, Locus Coeruleus, Raphe Nuclei, Nucleus Tractus Solitarius, Dorsal Vagal Nucleus, Spinal Cord

Central OT Projections

B.B. McEwen, 2004

Central OT Projections Prevailing view is that separate populations of

OT neurons project to posterior pituitary and to CNS structures

SON and PVN magnocellular neurons project to the posterior pituitary

PVN parvocellular neurons are the source of centrally projecting fibers

Evidence that central and peripheral OT release can be dissociated

Dendritic Release of OT Peptidergic neurons can release peptide from

entire cell surface, and peptide can diffuse long distances due to longer CNS half-life

OT can be released by dendrites independent of neuronal firing

Dendritically-released OT can have local autocrine/paracrine actions, or can diffuse to distant brain sites

Ludwig & Leng, 2006

Differential Regulation of Dendritic and Axonal OT Secretion

Ludwig & Leng, 2006

Role of OT in Stress and Emotion

A variety of stressors and stress paradigms result in increased plasma OT: Noxious Stimuli (e.g., footshock) Shaker Stress Forced Swimming Immobilization Stress Fear Conditioning

And increased secretion of CNS OT: Forced Swimming (increased PVN & SON OT) Social Defeat (increased SON OT) Shaker Stress (increased PVN OT)

Role of OT in Stress Given that OT is released in brain as a

function of various types of stress, what is the significance of this local hypothalamic release?

Since hypothalamic OT is released at the same time as activation of the HPA axis, does brain OT regulate HPA function?

Neumann, 2008

Central OT Regulation of HPA Axis

i.c.v. OT infusions in rats reduces plasma corticosterone release to wide variety of physical, emotional and pharmacological stressors in both male and female rodents

Suggests OT can modulate HPA axis, probably via inhibition of CRH in PVN

Elevated brain OT may serve to buffer the organism from stress

Neumann, 2008

Central OT Regulation of Anxiety-Related Behavior

i.c.v. administration of OT in both female and male rats exerted an anxiolytic effect (assessed by elevated plus maze)

These effects were localized to the central nucleus of the amygdala and PVN

Over expression of amygdaloid OTR in virgin female rats reduced anxiety-related behavior compared to control virgin females

Neumann, 2008

Stress Responses and Anxiety Related Behavior in OTKO Mice

Deletion of OT gene in females led to increased anxiety-related behavior and elevated plasma corticosterone during anticipation of shaker stress

In OTKO males, elevated corticosterone was seen following overnight food and water deprivation

Both OTKO and OTRKO mice exhibited heightened aggression compared to wild type mice

Amico et al., 2008

Central OT, Stress and Emotion

Increased central OT leads to reduced anxiety and calmness accompanied by blunted plasma glucocorticoid responses

It appears that the OT system is activated in response to particular stressors, and serves to attenuate both the physiological stress response and the emotional component of the response

Role of OT in Inflammation OT shown to reduce inflammation and oxidative stress

in several models: Carrageenan-Induced Inflammation (hindpaw) Wound Healing/Burns Sepsis Induction Colonic Inflammation Renal Injury

Suggests that OT is an endogenous anti-inflammatory and anti-oxidant molecule

Does OT play a role in the attenuation of disease via anti-oxidant and anti-inflammatory mechanisms?

Behavior & Atherosclerosis

In our lab, we sought to examine the influence of social environment and emotional behavior on the progression of atherosclerosis

Developed an animal model of disease that would allow us to study more easily the pathophysiological mechanisms in atherosclerosis

Watanabe Heritable Hyperlipidemic Rabbit (WHHL)

Model for human familial hypercholesterolemia Spontaneous genetic mutation in LDL receptor

synthesis Extremely high plasma lipids from birth Aortic atherosclerosis begins at 2 months, severe in

all animals by 7 months, CHD develops by 8-10 months, death occurs after 1 year

Since disease occurs spontaneously, can examine factors that slow the progression of disease as well as factors that advance disease progression

Social Environment and Progression of Atherosclerosis in the WHHL

(McCabe et al., 2002, Circulation)

33 male WHHLs, 3 months of age Assigned to one of 3 social conditions:

Unstable (paired with unfamiliar rabbit 4hrs/day, pairing rearranged each week)

Stable (paired with littermate 4hrs/day, pairing maintained throughout study)

Individually-caged (housed alone, no contact with other animals)

Study ran from 3 to 7 months of age

Area of Atherosclerosis as a Function of Social Environment

0

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Unstable Stable Individually-caged

Are

a o

f A

the

ros

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ros

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mm

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*

* p<.025

Study Conclusions

Stable social environment, characterized by increased affiliative behavior and decreased agonistic behavior, slows the progression of atherosclerosis by approximately 50% in animals genetically predisposed to disease

Group differences in disease could not be explained by differences in lipids, glucocorticoids, or gonadal steroids

Is disease attenuation in the Stable group related to oxytocin’s antioxidant and antiflammatory effects on vascular tissue?

Pathophysiology of Atherosclerosis

From Ross, R., NEJM, 1999

LDLLDL

LDLLDL

EndotheliumEndothelium

Vessel LumenVessel LumenMonocyteMonocyte

MacrophageMacrophage

AdhesionAdhesionMoleculesMolecules

Foam CellsFoam Cells

IntimaIntima

Oxidized Oxidized LDLLDLCytokinesCytokines

Further InflammationFurther InflammationCell ProliferationCell Proliferation

Matrix DegradationMatrix Degradation

CytokinesCytokinesGrowth FactorsGrowth Factors

MetalloproteinasesMetalloproteinases

MCP-1MCP-1MCP-1MCP-1NAD(P)HNAD(P)HOxidaseOxidase

Pathophysiological Processes in Atherosclerosis

INFLAMMATIONINFLAMMATIONOXIDATIVE STRESSOXIDATIVE STRESS

Adapted from P. Barter, Lipids Online

Oxytocin and Vascular Cells:In Vitro studies

(Szeto et al., Am. J. Physiol.: Endocr. Metab., 2008)

We have cultured human aortic endothelial cells, monocytes, macrophages and vascular smooth muscle cells

Incubated cells with physiological concentrations of oxytocin

Assessed the influence of oxytocin on oxidative stress (via NAD(P)H oxidase activity) and inflammation (via cytokine secretion)

Oxytocin Receptor Protein Expression

Oxytocin Inhibits Vascular Oxidative Stress in

Monocytes & Macrophages

Time (minutes)

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THP-1 Monocytes THP-1 Macrophages

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NADPH Oxidase in Aortic Endothelial & Aortic Smooth Muscle Cells

Time (minutes)

0 1 2 3 4 5 6 7 8

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Aortic Endothelial Cells Aortic Smooth Muscle Cells

OT Attenuates IL-6 Secretion from Vascular Cells in vitro

Aortic Endothelial Cells

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Control 10 pM OT 100 pM OT

**p<0.01; ***p<0.001

IL-6

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** p<.01, ***p<.001

Chronic OT Infusion in WHHL Rabbits

OT (n=14) or saline (n=14) infused chronically via osmotic minipumps for 4 months beginning at 3 months of age

Rabbits housed individually for entire study

Aortas and Adipose tissue removed at endpoint. Ex vivo secretion of IL-6 by adipocytes measured

Vehicle

Oxytocin

* **

Chronic in vivo OT infusion in WHHL rabbits

Oxytocin and Fat Cells Adipose tissue is a major source of

proinflammatory cytokines Increased adiposity is a risk factor for a

number of inflammatory diseases, including cardiovascular disease

Does oxytocin influence the secretion of proinflammatory cytokines from adipocytes?

Epididymal Fat IL-6 mRNA Expression in WHHL Rabbits

In Vivo OT infusion in ApoE Knockout Mice (Nation et al., Psychosom Med, In Press)

Recently, we demonstrated that chronic OT infusions significantly attenuated atherosclerosis in a transgenic murine model, the ApoE knockout mouse

OT infused mice exhibited less disease in the thoracic aorta than vehicle infused animals

Ex vivo IL-6 release from adipocytes was attenuated in OT infused mice.

Conclusions

Stable social environment, characterized by increased affiliative behavior & less agonistic behavior, slows the progression of atherosclerosis in animals genetically predisposed to disease

Vascular cells, macrophages, and adipocytes express oxytocin receptors

Oxytocin inhibits oxidative stress in cultured vascular cells, and inflammatory cytokine release in macrophages and adipocytes

Conclusions

In vivo infusion of OT slows the progression of atherosclerosis in a site-specific manner

It is proposed that the beneficial effects of a prosocial environment on disease progression may be mediated through the direct effect of peripheral oxytocin on pathophysiological mechanisms occuring in vascular wall

Collaborators

University of Miami Department of Psychology: Angela Szeto, Daniel Nation, Larry Brooks, Crystal

Noller, Maria Rossetti, Julie Gonzales, Jim Paredes, Maria Llabre, & Neil Schneiderman

University of Miami Medical School: Armando Mendez, Julie Zaias, Ron Goldberg

Eehscience LLC, Pickerington, OH Edward Herderick