Dissertation Proposal

The Effects of Ethanol and Ethanol Metabolites on

Locomotor Activation and Ataxia: Focus on the

Substantia Nigra pars Reticulata

Maria Arizzi, M.A.Dissertation Proposal

Biphasic Effects

• Ethanol is typically classified as a sedative hypnotic

• Many studies have shown activating as well as depressant effects after ethanol administration

Biphasic Effect in Mice and Humans• Biphasic effects first shown in mice

Read et al. 1960

• Many studies since have confirmed this finding

• Recent research in humans has also shown biphasic effects of ethanol– Self-report – BAES Earleywine and colleagues

– Biological measurements Davidson et al. 1997, 2002, Grassi et al. 1989, Higgins et al. 1993, Inder et al. 1995, Rush et al. 1993

Biphasic Effects in Rats

• Only sedative effects have been seen in rats via peripheral administration (i.p. or oral) of ethanol Bass et al. 1979, Duncan et al. 2000, George et al. 1990, Gingras et al. 1996, Little et al. 1996, Moore et al. 1993, Petry et al. 1998, Pohorecky 1977

• Recent work using intraventricular administration has shown a biphasic effect of ethanol in rats

Effects of Ethanol, Acetaldehyde, and Acetate on Locomotor Activity in Rats: Open Field

• In the proposed set of experiments the focus will be on the activating effects low doses ethanol

0.0 0.7 1.4 2.8

0

5

10

15

20

25

30

35

40

45

50

55

60

ETHANOL DOSE (moles/l)

MO

TO

R A

CT

IVIT

Y (

sc

ore

s i

n 1

0 m

inu

tes

)*

** **

Ethanol infused into lateral ventricles: locomotor activity First 10 minutes stabilimeter

Ethanol Metabolism

Peripheral MetabolismEthanol

ADH, CYP 450, CAT

Acetaldehyde

ALDH

Acetate

Brain MetabolismEthanol

CAT

Acetaldehyde

ALDH

Acetate

Are the metabolites active?

Effects of Ethanol, Acetaldehyde, and Acetate on Locomotor Activity in Rats: Open Field

0 0.7 1.4 2.80

20

40

60

80

100

120

ACETALDEHYDE DOSE (mol)

****

*

MO

TO

R A

CT

IVIT

Y (

cou

nts

in 1

0 m

inu

tes)

ETHANOL DOSE (mol)

Ethanol and Acetaldehyde infused in to the lateral ventricles: First 10 minutes, stabilimeter

Working Hypotheses

• Acetaldehyde increases behavior– Increases open field and stabilimeter

activity– Increases operant behavior on low rate

schedules

• Acetate decreases behavior– Decreases open field activity– Decreased operant on high rate

schedules

Dose (mol)

0 4.4 8.8 17.6

Mea

n L

ever

Pre

sses

200

400

600

800

1000

1200

1400

1600

1800

2000

EthanolAcetaldehydeAcetate

*

*

FR5 high doses

What are the brain areas that mediate the activating effects of ethanol and

acetaldehyde?

•VTA?•NAcc?•CPU?•SNr?

SNr and Motor Behavior

• Muscle rigidity Crocker 1997

• Lever pressing Correa et al. submitted, Trevitt et al. 2001

• Tremor Finn et al. 1997, Mayorga et al. 1999

• Catalepsy Scheel-Krueger et al. 1977

• Circling Scheel-Krueger et al. 1977

• Locomotion Abraini et al. 1999, Scheel-Krueger et al. 1977, 1981, Trevitt et al. 2002

SNc

STRIATUM SNr

DA

GABA

SNr

STRIATUM

STN

SuperiorColliculus

ReticularFormation

BrainstemMotor

MotorThalamus

SNr, GABA, and Motor Behavior

• GABA agonists and antagonists have their most potent effect on locomotion when infused into the SNr

• Dopaminergic stimulation in the SNr increases locomotion and increases GABA release in the SNr

• Locomotor effect attenuated by infusion of GABAA antagonist

Trevitt et al. 2002

Ethanol and GABA

• Modulation of GABA transmission

• Facilitation of GABAA receptor function

Why SNr?• GABA manipulations in SNr effect

locomotor behavior• Ethanol works through the GABA

receptor• SNr is a part of the basal ganglia

circuitry important for the modulation of locomotion and motor function

• SN has relatively high density of catalase and is therefore a possible place for acetaldehyde action

Focus on the SNr

SNc SNr

List of ExperimentsExperiment Task Drug

Experiment 1 Locomotor Activity Ethanol

Experiment 2 Locomotor Activity Ethanol, dorsal site

Experiment 3 Locomotor Activity Acetaldehyde

Experiment 4 Locomotor Activity Acetate

Experiment 5 Locomotor Activity Ethanol, sodium azide

Experiment 6 Locomotor Activity Acetaldehyde, sodium azide

Experiment 7 Ataxia Ethanol

Experiment 8 Ataxia Acetaldehyde

Experiment 9 Ataxia Acetate











Preliminary data

Ethanol infused in SNr: Locomotor ActivitySecond 5 minutes (bin 6-10)

Dose (mol)

0 0.175 0.35 0.7 1.4 2.8

Mea

n ac

tivity

cou

nts

(bin

6-1

0)

10

15

20

25

30

35

40

Ethanol

*

* denotes significant difference p<.01 Preliminary data

Preliminary data











Sedation Scale• 4 – awake, active

– Engaged in locomotion, rearing, or head movements

• 3 – awake, inactive– Eyes fully open, head up, no locomotion or rearing, normal

posture

• 2 – moderate sedation– Eyes partly closed, head somewhat down

• 1 – heavy sedation– Eyes mostly closed, head mostly or completely down,

flattened posture, lack of normal limb placement

• 0 – asleep– Eyes fully closed, body relaxed, loss of righting reflexSalamone et al. 1996

Hypotheses• Ethanol infused into the SNr will have

an activating effect on locomotor behavior at low doses

• Acetaldehyde infused into the SNr will also produce an activating effect, and will be more potent than ethanol

• Acetate infused into the SNr will not increase motor activity and may produce a suppressive effect on activity

Hypotheses (continued)

• The behavioral activating effect of ethanol infused in the SNr can be blocked with peripheral administration of a catalase inhibitor. The catalase inhibitor will have no effect on acetaldehyde-induced behavioral activation

Hypotheses (continued)

• Low doses of ethanol and acetaldehyde infused into the SNr will not produce ataxia

• Of all three compounds studied, acetate is the most likely produce ataxia in the dose range tested

• Locomotor activity and ataxia are dissociable and are being produced through 2 different mechanisms

The results of the proposed experiments will further elucidate the brain mechanisms and biochemical pathways through which ethanol exerts some of its behavioral effects.

Effects on neurotransmission

GABA. Like benzodiazepines, barbiturates and anesthetics, acute ethanol enhances GABA’s influence on GABAA receptor thus facilitating Cl- influx. (10-50 mM). This process seems to be mediated by the phosphorylation induced by PKC (protein kinase C). Chronic ethanol administration produces the opposite effects on GABAA receptors and reduces the synthesis of one of protein subunits of this receptor.

Glutamate. Acute administration inhibits NMDA receptor/cation channel complex (20-50 mM). This inhibition interferes with excitotoxicity in cortex, and with LTP in the hippocampus. Chronic administration increases NMDA receptor numbers.

DA. Acute and Chronic alcohol administration induces

dopamine release in the N. Accumbens.

5-HT. Acute ethanol induces the release of serotonin

and facilitates the cation-5-HT3-gated channel (10-50 mM). Chronic alcohol increases the number of 5-HT2.


Endogenous Opioids. Acute alcohol

increases endorphin and enkephalin expression and

release. Chronic alcohol produces the opposite

effects.

Adenosine. Acute alcohol increases extracellular

adenosine by inhibiting adenosine uptake, and

increases synthesis of adenosine. The activation of

the A2 receptor increases intracellular cAMP. Chronic

alcohol produces the opposite effects.


Evidence of central ethanol metabolism by Catalase.

Catalase is widespread in brain, and also is localized in several discrete brain nuclei.

In vivo and in vitro administration of catalase inhibitors blocks acetaldehyde formation in different brain nuclei. In contrast, when H2O2 concentration is increased, acetaldehyde fomation also increases.

Acatalasemic mice mutants have less ethanol-induced locomotor activity, more ethanol-induced sleep, and higher ethanol consumption.

The pharmacological inhibition or potentiation of catalase modifies some acute ethanol-induced effects as well as the ingestion of ethanol.

Dissertation Proposal

Documents

ethanol ataxia experiment

acetaldehyde ataxia

ethanol administration

ethanol metabolism

ethanol metabolites

biphasic effects ethanol

activating effects of

snr locomotor effect