Plant and brain cannabinoids:major players in physiology.

Raphael Mechoulam

Beit Dagan, January 2007 LOH 2003

Gan-zi-gun-nu – the drug that takes away the mindAzallu – hand of ghost, poison of all limbs (neurological diseases?)Qunnabu – used in religious rites

Pliny, the Elder (79 AD): The roots boiled in water ease cramped joints, gout too and similar violent pain.

Dioscorides (90 AD): The sodden root when placed on inflammations sooths them, eliminates edema and disperses obdurate matter above inflammed joints.

LOH 2003

For the relief of certain kinds of pain, I believe, there is no more useful medicinethan Cannabis within our reach.

J. Russell Reynolds, Archives of Medicine, Vol 2, 154, 1859

-tetrahydrocannabinol (-THC) (Gaoni and Mechoulam, 1964)

CH3

O

OH

C5H11

cannabidiol (CBD)(Mechoulam and Shvo, 1963)

HO C5H11

OH

CH3

CH3

cannabigerol (CBG)(Gaoni and Mechoulam, 1964)

HO

OH

C5H11

cannabichromene (CBC) (Claussen et al., 1966;Mechoulam and Gaoni, 1966)

O C5H11

OH

Representative natural cannabinoids

CH3

cannabinol (CBN)(Adams et al., 1940)

O

OH

C5H11

O C5H11

OH

cannabicyclol (CBL)(Crombie et al., 1968)

LOH 2003

Gaoni and Mechoulam: J.Amer.Chem.Soc. 86, 1646 (1964)

cannabidiol (CBD)

HO C5H11

OH

CH3

O

OH

CH 3

9-tetrahydrocannabinol (9-THC)

Mechoulam and Shvo: Tetrahedron 19,2073 (1963)

Brain regions in which cannabinoid receptors are abundant

Reward pathwayNucleus accumbens

Link between cerebral hemispheresIntrabulbar anterior commissure

Higher cognitive functionCerebral cortex, especially cingulate, frontal, and parietal regions

Learning and memory, stressHippocampus

Body-movement coordinationCerebellum

Putamen

Globus pallidus

Enteropeduncular nucleus

Substantia nigra pars reticulata

Movement controlBasal ganglia

LOH 2004

anandamide

-tetrahydrocannabinol

-THC)

O

OH

CH3

OC

O

OH

OH

2-arachidonoyl glycerol (2-AG)

N

OC

H

OH

Binding to cannabinoid receptors

Ligand CB1

(nM)

CB2

(nM)

GPR55

EC50 (nM)

9-THC 6 0.4 8

cannabidiol > 30000 > 30000 445 *

anandamide 31 27 18

2-arachidonoylglycerol 519 618 3

palmitoylethanolamide > 30000 19800 4

HU-210 0.2 0.5 26

G G

AC AC

Calciumentryblocked

[Ca2+]Decreasedrelease ofneurotransmitters

Cannabinoidreceptor

Agonists (e.g. THC,anandamide)

Ca2+

(-)(-)

K+

PotassiumPotassiumchannelschannelsopenopen

[cAMP]

(-)K+

Science (STKE, 23 April, 2002)

What do endocannabinoids do?

“Relax, eat, sleep, forget and protect”

Di Marzo, 1998

Physiological systems and conditions affected by endocannabinds (a partial list)

Anxiety Inflammation

Appetite/feeding Memory Blood pressure Mood

Bone formation Movement Cerebral blood flow Neuroprotection Digestive system Pain Emesis and nausea Reproduction Immune system Stress

Neuroprotection

control 15 min 1 h 4 h 8 h 24 h 0

10

20

30

40

50

60

70

80

90

100

110

120

Levels of 2-AG in mouse brain after CHI

Anova with Tukey post-test:P<0.0001, F=36.01

-P<0.01 vs. control

-P<0.05 vs. control

Time after CHI

con

cen

trat

ion

of

2-A

G(n

M/g

r)

-P<0.001 vs. control

Nature 413, 527 (2001)

sham CHI 0.1 5 10 (mg/kg)

76.5

77.5

78.5

79.5

80.5

81.5

Effect of exogenous 2-AG on edema after CHI

-P<0.05 vs.CHI

0.0

0.5

1.0

1.5

2.0

2.5

3.0

NS

S (

imp

rove

men

t)

CHI 0.1 5 10 (mg/kg)

Effect of exogenous 2-AG on recovery after CHI

w

ate

r

*

2-AG Reduces Infarct Volume 24 h After CHI2-AG Reduces Infarct Volume 24 h After CHI

control 2-AG (5 mg/kg)0.0

2.5

5.0

7.5

10.0

12.5

15.0control2-AG 5 mg/kg

*

n=9 n=7

unpaired t-test, P=0.0 3

infa

rct

vo

lum

e(%

)

2-AG

control

glutamatecytokines, ROS

cerebralischemia

neuronal & glial cell death

Brain injury

Vasoconstrictors(e.g. ET-1, Thromboxane)

2-AG

cerebroprotection

Regulation of vasodilation

Corticosteroids

Androgens

Estrogens

AldosteroneProgesteron

Costicosteroids

Digitalis

HO

Plasma MembranePlasma Membrane

CO

O

CO

O

OP

O O

ONH2-

phosphatidylethanolamine

R1

R2

CO

O

CO

O

OP

O O

ON-

N-arachidonoylphosphatidylethanolamine

R1

R2

CO

H

HON

CO

H

anandamide

CO

O

CO

O

OP

O O

ONH2-

R1

R2

COOH

phosphatidylserine

CO

O

CO

O

OP

O O

ON-

N-arachidonoylphosphatidylserine

R1

R2

CO

H

HON

CO

H

arachidonoylserine

COOH

COOH

CN

OHHO

CN

OH

O HCOOH

anandamide

arachidonoyl-serine

anandamide

OC

O

OH

OH

2-arachidonoyl glycerol (2-AG)

N

OC

H

OH

HO

COOH

N

OC

H

arachidonoyl serine

Inflammation

0 0.05 0.1 1 5 10 50

0

50

100

150

200T

NF

pro

du

cti

on

(S

50

un

its

)

Inhibition of TNF production by 2-AGIn vitro

2-AG (mg/ml) LOH 2004

Eur.J.Pharmacol. 406, R5-R7 (2000)

0

500

1000

1500

2000

LPS LPS + 2-AG LPS + 2-AG + +2-LG + 2-PG

TN

F

in

ser

um

(S

50 u

nit

s)Inhibition of TNF production by 2-AG in mice

In vivo

LOH 2004

0 2 4 60

100

200

300

400

500

CBD (g/ml)(macrophages)

TN

F

tite

r (S

50)

0 2 4 60

1

2

3

4

5

6

CBD (g/ml)(macrophages)

NO

(n

M)

0

500

1000

1500

2000

zymosan zymosan +CBD

zymosan+ CBD (- 1 h)

ROI(granulocytes)

chem

ilum

inis

cenc

e pe

ak

Synovium is the most critical site of cytokineproduction in arthritis. Synovial cells from arthritic mice spontaneously produce large amounts of TNF when cultured in vitro. Cells from arthritic mice which had been treated with CBD produced significantly less (50%) TNF.

LOH 2003

Cannabidiol (i.p.), acute

0 1 3 5 7 9 110

1

2

3

4

5

control2.5 mg/kg5 mg/kg10 mg/kg20 mg/kg

Days after onset of arthritis

Cli

nic

al s

core

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Appetite and feeding

0 14 28 42

-0.4

-0.3

-0.2

-0.1

-0.0

0.1

0.2

0.3

Day

Mea

n w

eig

ht

chan

ge

(kg

)

Dronabinol

Placebo

Mean change in weight from baseline in patients with AIDS treated with dronabinol (THC) and placebo (Beal et al., 1995)

0 1 2 3 4 5 6 7 83

4

5

6

7

anandamide (0.001 mg/kg i.p.)vehicle

Days

gram

/mou

se/d

ay

Effect of anandamide on food intake (16 mouse in each group).

LOH 2004

LOH 2004

Cl

NN

N N

O

Cl

Cl

H

SR141716A

Memory extinction

d1 d2 d3 d60

10

20

30

40

50P

er c

ent

free

zin

g

(60

s t

one)

CS CS CS CSCo

Experimental day

*****

***

………

……

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The endogenous cannabinoid system could represent a therapeutic target for the treatment of diseases associated with inappropriate retention of aversivememories or inadequate responses to aversive situations, such as post-traumaticstress disorders, phobias, and certainforms of chronic pain.

LOH 2004

Cannabinoids and anxiety.

1. In animal models as well as in limited clinical trials cannabidiol has been shown to lower anxiety. Mechanism unknown. 2. Blocking of anandamide hydrolysis in vivo (rats) enhances anandamide levels in brain and lowers anxiety. This effect is blocked by endocannabinoid antagonists.

Cannabinoids and schizophrenia.

1. Large doses of cannabis (particularly if they lack cannabidiol) may cause a psychotic state resembling psychosis. 2. In schizophrenic patients cannabis use may worsen positive symptoms. 3. Heavy use of cannabis may precipitate schizophrenia in susceptible individuals. Frequent cannabis use was associated with 6 fold increase of in high risk individuals (family history). 4. Increase in density of CB1 receptors in several brain areas in postmortem schizophrenic brains. Two fold increase of anandamide levels in CSF of patients.

Cannabidiol treatment of positive schizophrenia.

High doses of the non psychoactive Cannabis constituent cannabidiol have shown effectiveness in the treatment of schizophrenics (lowering of positive effects). Mechanism – unknown.

Endocannabinoids and depression.

Although at low doses cannabis may enhance mood it is not antidepressive.

A selective inhibitor of the enzyme which causes endocannabinoid hydrolysis (in vivo) exerts potent anti-depressant effects in the rat forced swim test.

Hepatic encephalopathy.

Hepatic encephalopathy (HE) is a neuropsychiatric syndrome due to liver disease. It is the feature that defines prognosis in acute liver injury.

1. In a mouse model of HE (produced by the toxin thioacetamide) 2-AG levels are enhanced in the brain. 2. Administration of either 2-AG or the specific CB2 agonist HU-308 improves a neurological score, activity, cognitive function. 3. A specific CB2 receptor antagonist blocks these effects.

Alzheimer’s disease

Compared to currently approved drugs prescribed for the treatment of Alzheimer’s disease, THC is considerably superior inhibitor of A aggregation and thus cannabinoid molecules may directly impact the progression of this debilitating disease.(Eubanks et al., Mol. Pharmac. 2006)

Collaboration in Israel

Jerusalem

Prof. L. HanušProf. E. FrideDr. W. A. DevaneDr. A. BreuerDr. S. Ben-ShabatD. PanikashviliG. Milman N. KoganY. Maor

Jerusalem

Prof. E. ShohamiProf. R. GallilyProf. E. BerryProf. M. SchlesingerDr. Y. Avraham

Haifa

Prof. A. Mandelbaum

Rehovot

Prof. Z. Vogel

Tel Hashomer

Dr. S. AlmogDr. A. Gopher

LOH 2003

Collaboration abroad

AberdeenR. Pertwee

BonnM. KarsakA. Zimmer

BrnoA. Šulcová

GreeceC. Simeonidou

RichmondB. MartinA. H. Lichtman

CanadaL. A. Parker

BethesdaG. KunosM. Spatz

NapoliV. Di Marzo

RomeM. Maccarrone

SiberiaL. Maslov

LondonM. FeldmannA. M. MalfaitP. F. Sumariwalla LOH 2003