Are Cannabinoids Neurotoxic-12

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Are Cannabinoids Neurotoxic?

Introduction

Cannabis is the worlds most commonly used illicit drug (United States

Department of Health and Human Services, 2000). While the debate continues as to

whether cannabis laws should change, so does the debate on the extent of suggested

negative effects of cannabis use (Hall, 1997). The possible negative effect of cannabis

and its major psychoactive component Δ9

-tetrahydrocannabinol (THC), on the brain and

specifically neurotoxicity, is one of the many topics brought up in such debates. While

there is a large number of research articles investigating cannabinoid neurotoxicity, there

has only been one review dedicated to this field (Scallet, 1991). Since this review was

written, over a decade ago, the landscape of cannabinoid research has changed

significantly. The existence and nature of the CB1 and CB2 receptors has been widely

studied (Felder & Glass, 1998), on top of that there has been the discovery of anandamide

(Devane et al., 1992) (AEA), sn-2-arachidonylglycerol (Mechoulam et al., 1995) as well

as other endo-cannabimimetics (Lambert & Di Marzo, 1999). Therefore, it seems that a

new review may add to the field.

When discussion neurotoxicity, it is important to consider what is actually meant

by neurotoxicity. The Interagency Committee on Neurotoxicology (ICON) adopted the

definition that neurotoxicity is “any adverse effect on the structure or function of the

central and/or peripheral nervous system by a biological, chemical or physical agent and



may result from direct or indirect actions or reflect permanent or reversible changes in the

nervous system” (see Scallet, 1991).

While the ICON definition includes reversible changes, it is important to consider

that while a drug is still in an animal, it will still have an effect and even if this effect is

adverse, it is hardly reasonable to consider a drug neurotoxic if the adverse effect

disappears as soon as the drug is cleared from the system. Hence, before any parameters

are measured, it is important to have a “washout phase” where the animal is given no

drug. Indeed, this is especially important when examining THC, as its highly fat soluble

nature and active metabolites allows it to persist in the plasma for weeks (Grotenhermen,

2003). In humans, a long washout phase is important not only to exclude residual drug

actions, but also to allow any possible withdrawal effects to subside (Smith, 2002).

Many studies have claimed to show evidence for cannabinoid-induced

neurotoxicity on the basis that chronic administration of cannabinoids leads to altered

morphological or neurochemical parameters. In light of the definition of neurotoxicity

mentioned above, this is not sufficient, as changes need to be negative to be considered

neurotoxic. Although some researchers might suggest that any changes in the brain are

negative, this is obviously not the case. Consider the example of large-scale differences

seen between the brains of animals raised in standard and enriched environments (van

Praag et al., 2000). Here we have neurochemical and morphological changes that

correlate with positive changes in behaviour, so all neuronal changes are not bad. One

could then argue that there would be some effects that would be obviously good, like

neuronal proliferation, and some that would be obviously bad, such as cell death, but in

some instances the same effect can be seen in both toxic and beneficial situations. For



example, the proliferation of astrocytes is seen associated with Alzheimer’s disease

(Coria et al., 1993; Luth et al., 2003) as well as in the brains of animals raised in enriched

environments (Kempermann et al., 1997; Walsh et al., 1969). Therefore, with the

exception of cell death, unless there is data which shows negative behavioural changes

that are directly correlated with a physical (morphological or neurochemical) change,

then the physical change can not be considered as evidence for neurotoxicity.

A similar problem is raised when considering behavioural results. Just because a

behaviour is changed, does not mean that there has been a negative impact on an animal.

Indeed, the same change of behaviour can be seen as positive by one group of

experimenters and negative by another. For instance, increased exploratory behaviour in

an open field test is seen as a negative by someone investigating the hippocampal toxicity

of kainic or domoic acid (Strain & Tasker, 1991), but is seen as a positive by someone

examining the anxiolytic action of postnatal handling (Padoin et al., 2001). Fortunately,

in cannabinoids research, much of the data gathered is in regards to learning and memory,

and in most cases it can be agreed that a reduction in learning or memory is a negative

impact. But where other behaviour is studied the question of whether or not any given

effect is ‘negative’ needs to be considered.

The example used above shows us another caveat. If a lesion or neurotoxin brings

about a behavioural effect e.g. kainic acid induced increase in exploratory behaviour in

an open field test, it does not mean that an intervention which produces the same

behavioural effect produces the same neurotoxicity, such as postnatal handling.

Unfortunately, a researcher can not always look for neurological changes induced

by a drug and then check to see if there is a behavioural correlate. Therefore, we must



closely compare neurochemical/neurohistological experiments, which show a physical

change, with behavioural experiments, which show a behavioural change, in order to

make a conclusion about proposed neurotoxicity.

Neuronal Morphology

The classic first step when investigating whether or not a given chemical is

neurotoxic, is to give it to an animal and then investigate, using histological methods,

whether there has been any cell death or at least alteration of cellular morphology.

The first study into the effect that cannabinoids have on neuronal morphology was

Harper et al ., (1977). This study reported that rhesus monkeys exposed to the smoke of 1

to 3 2.6% THC cannabis cigarettes a day or 0.7mg/kg THC a day for 6 months showed

widening of the synaptic cleft, material in the synaptic cleft and nuclear inclusion bodies

after an 8-month washout phase. Unfortunately there were many methodological

problems with this study. There were no statistical analysis, the sample sizes were

extremely small, consisting of one active cannabis smoke treated animal, one IV treated

animal and the controls were two completely untreated monkeys and one monkey who

smoked alcohol extracted cannabis leaf which was presumed to be THC free. Indeed, the

apparatus in which the animals were placed in order to administer the cannabis smoke, a

procedure that most of the control animals were free of, seemed to be so constrictive and

unnatural, that it could be a serious source of bias.

Scallet et al., (1990) repeated this experiment, with larger sample numbers and

showed that after a wash out phase of 7 months, rhesus monkeys who were exposed to

the smoke of a single cannabis cigarette containing 2.6% THC, every day for 12 months,



showed no statistically significant changes in synaptic characteristics, neuronal size, the

number of apical or basilar dendrites or the number or length of dendritic branches, in

comparison to monkeys who were either non-exposed or smoked ethanol-extracted

cannabis. Andrews et al ., (1989, as cited by Scallet et al ., 1991) also found no

neurohistological changes after dosing moneys IV with either 0.1 or 1mg/kg of THC for

90 days.

Scallet et al ., (1987) examined the hippocampal morphology of rats that had been

dosed orally with THC 5 days a week for 90 days with 20mg/kg Monday to Thursday and

60mg/kg on Friday. After a 7 month washout phase, the THC treated animals were found

to have a significant decrease in the cross sectional area of neurons and their nuclei, as

well as a decrease in the synaptic density in the CA3 region of the hippocampus, in

comparison to vehicle treated controls. Cerebellar Purkinje cells were unaffected. A

second group of animals was dosed with either 10mg/kg or 20mg/kg THC, 5 days a week

for 90 days. After an 8-week washout phase Golgi impregnation showed a reduction in

the length of the outermost branchlets of CA3 neurons. Interestingly, in the second group

of rats, there was no reduction in the size of neurons or synaptic density.

Landfield et al ., (1988) treated rats 5 days a week for 8 months, with either

4mg/kg or 8mg/kg of THC orally. The 8mg/kg group originally received 10mg/kg but

this dose was lowered during the first month because the animals found it aversive.

Directly after the last dose of THC the CA1 region of the hippocampi from the high dose

animals were shown to have a decreased number of pyramidal cell per area of section.

Whether this reduction in pyramidal cell density was due to decreased cell volume or



actual loss of cells was not explored. It was also shown that there was an increase in dark

membranous inclusions in hippocampal astrocytes in THC treated animals.

Chan et al ., (1996) dosed rats orally with 0-500mg/kg THC 5 times a day, for

various time periods. Some of the rats suffered convulsions from such high doses of THC

and so at 9 and 15 months, the neurohistology of only the convulsive rats was

investigated. It was reported that “no treatment-related neuropathological changes were

observed in any tissue evaluated by step section”. Unfortunately, there is no report on the

number of rats in this sample, or on the specific parameters measured.

Lawston et al ., (2000) showed that rats that were subcutaneously injected twice

daily with 2mg/kg of the synthetic CB1 receptor agonist WIN 55,212-2 for 21 days,

exhibited cellular alterations in the hippocampus. Using MAP-2 antibodies, which act as

a dendritic marker it was shown that there was increased staining in the subiculum and

CA3 region of the hippocampus, as well as increased staining in the lower blade of the

dentate gyrus in comparison to vehicle treated controls. It was also shown that there was

a decrease in MAP-2 staining in the CA1 region of the hippocampus and an increase in

cresyl violet staining in the lower blade of the dentate gyrus. The authors also stated that

the dendrites of CA1 neurons in treated animals appeared as disjointed segments, which

seemed twisted or broken with a beaded aspect, rather than continuos smooth, structures,

as was seen in their controls animals. Although this is an intriguing claim, there are no

statistics associated with it, and so the definitiveness of this claim can not be verified.



Neurochemistry

On top of neurohistological methods, another technique used to investigate the

potential neurotoxicity of a chemical, is to look for a neurochemical change associated

with it. If a chemical causes an alteration in a neurochemical system, and this is

associated with a negative behavioural change, then this fits into the ICON definition of

neurotoxicity. In neurochemical studies one must be very careful to allow for an adequate

washout phase, to make sure that any potential change is not a transient one that will

dissipate as the cannabinoids are cleared from the body.

Walters and Carr (1986) were the first to investigate the neurochemical alterations

induced by chronic cannabinoid administration after a significant washout phase. In this

experiment, pregnant rats were dosed with a cannabis extract daily so that they received

20mg/kg THC, for 20 days prior to impregnation until 20 days after giving birth, when

the offspring were weaned, which means approximately 60 days of dosage. It was

reported that at both 10 and 20 days after birth, the offspring had a significant decrease in

the Bmax of D2 receptors in striatal tissue samples in comparison to vehicle treated

controls. It was also reported that at day 20 after birth, the offspring had a significant

decrease in the K d of α1 receptors. At days 40 and 60 after birth, the offspring showed no

significant changes in Bmax or K d of either receptor studied. Tyrosine hydroxylase activity

was also studied, and in comparison to the controls it was shown to be significantly

decreased in the striatum at days 20 and 40 after birth, and in the cortex at day 40 after

birth. It is interesting to note that the cannabinoid treated animal gained significantly less

weight during pregnancy and that their offspring significantly weighed significantly less

and had lighter brains than the controls. The time points when there was a significant



alteration of neurochemical parameters were nearly exclusively during times when the

animals were still receiving cannabinoids (apart from one tyrosine hydroxylase activity

recording), as they were still suckling at mothers who were being treated with

cannabinoid extract. All recorded neurochemical parameters had returned to normal after

being free of cannabinoids for 20 days. This could point to the fact that any alteration in

receptor binding characteristics were due to the direct effect of cannabinoids, and did not

reflect any neurotoxicity, though, the lowered brain weight induced by cannabinoid

treatment could refute that.

The experiment of Walters and Carr (1986) was repeated, though this time instead

of pregnant rats receiving cannabis extract, they were exposed to either THC (10mg/kg),

Δ8-tetrahydrocannabinol (Δ8-THC) (1mg/kg) or cannabidiol (CBD) (10mg/kg) (Walters

& Carr, 1988). The same neurochemical parameters where recorded and the rats were

dosed in the same fashion. It was reported that CBD was the only cannabinoid to alter D 2

receptor K d at any time point and no cannabinoid altered D2 receptor Bmax. CBD was

reported to significantly increase D2 K d in offspring at 10 and 20 days after birth in

comparison to control. It was reported that THC and Δ8-THC significantly increased α1

receptor Bmax at day 20 after birth, while CBD significantly increased α 1 receptor K d at

day 10 in comparison to control. It was reported that tyrosine hydroxylase activity was

significantly decreased at day 60 after birth by Δ8-THC and CBD.

When comparing these two experiments, it is interesting to note that although

they used the exact same methods in regards to everything apart from the fact that

Walters and Carr (1986) used a cannabis extract, while Walters and Carr (1988) used

separate cannabinoids, their results completely disagree. This indicates that either a



cannabinoid apart from the ones used by Walters and Carr (1988) are responsible for the

results of Walters and Carr (1986), that cannabinoids administer together have nearly the

opposite effect from when they are administered separately, or that there was a

methodological error in one of these experiments.

Ali et al ., (1989) also investigated what neurochemical effects chronic THC

administration produced in the rat, after a significant washout phase. Rats were dosed

orally for 90 days, 5 days a week, with 0, 10 or 20mg/kg THC then sacrificed either

directly after the last dose or after a 2-week washout phase. It was reported that when

rats were sacrificed directly after the last dosing, neither the high or low dose groups had

any significant alteration in the Bmax of GABA, muscarinic acetylcholine, dopaminergic,

or mu, kappa and delta opioid receptors in the hippocampus in comparison to the controls

or in dihydroxyphenylacetic acid (DOPAC), 5-HT or 5-hydroxyindoleacetic acid (5-

HIAA) concentration in the hypothalamus, septum or caudate nucleus. In rats sacrificed

after a 2-week washout phase there was found to be a significant decrease in the Bmax of

GABA receptors in the high dose group, though no other changes were found. In order to

repeat these results, and to see whether chronic THC treatment had any effect on the

properties of sigma opioid receptors or various allosteric GABAA receptor sites, a second

experiment was designed. In this experiment, rats were treated in the same fashion as the

first experiment with the addition of a fourth dosage group that received 20mg/kg

Mondays-Thursday and 60mg/kg on Friday. In this experiment, binding was only

investigated after a 2-week washout phase. It was reported that there was no significant

alteration in the Bmax of sigma receptors, GABA receptors, or any of the GABAA

allosteric sites at any dosage level, in disagreement with their earlier results. In the end,



these results are inconclusive, and no other study has looked into the effect of chronic

cannabinoid treatment of the GABA receptor since.

Ali et al ., (1991) examined the effects of chronic THC or cannabinoid treatment

in rats and the rhesus monkey respectively. Rats were dosed with orally with 10 or

20mg/kg of THC 5 days a week, for 90 days and sacrificed 2 hours or 2 months after the

last dose. It was reported that at either 2 hours or 2 months, neither of the doses produced

a significant effect on the Bmax of muscarinic acetylcholine receptors in comparison to

vehicle treated controls. Rhesus monkeys were treated for either 2 or 7 days a week for 1

year with the smoke of 1 2.6% cannabis cigarette, the smoked of an ethanol extracted

cannabis cigarette or no smoke and then sacrificed 7 months after the last treatment. It

was reported that there were no statistically significant dose-related changes in dopamine,

DOPAC, serotonin or 5-HIAA concentration, in either the caudate nucleus or

hypothalamus.

Westlake et al ., (1991) investigated whether chronic treatment with THC altered

the Bmax or K i of the cannabinoid receptor in either rats or rhesus monkey. Rats were

dosed with 0, 10 or 20mg/kg 5 days a week or 20mg/kg Monday to Thursday and

60mg/kg Friday for 90 days and sacrificed 60 days after the last dose. It was reported that

there were no significant changes induced by any dose in either the Bmax or the K i of the

cannabinoid receptor in the cortex, striatum, cerebellum, hippocampus or brainstem in

comparison to the control. Rhesus monkeys were exposed to the smoke of either ethanol

extract cannabis or 2.6% THC cannabis, 7 days a week for a year and then sacrificed 7

months after the last exposure. It was reported that there were no significant changes in



the Bmax or the K i of the cannabinoid receptor in the cortex, caudate nucleus or the

cerebellum in comparison to the control.

Animal Behavioural

As mentioned earlier if a chemical is to be considered neurotoxic it must produce

not only some measurable change in neuroanatomy or neurochemistry, but it must

produce a negative behavioural change. This behavioural change must also continue after

the drug has left the body.

The first experiment that investigated what behavioural effects chronic

cannabinoid treatment may have, after a significant wash out phase was Fehr et al .,

(1976). The rats were dosed orally with an ethanolic cannabis extract containing 10mg/kg

THC daily for 30, 60 or 90 days or 20mg/kg THC for 180 days, and then after a 1-month

washout phase. After the washout phase the animals ability in a moving belt task and the

Hebb-Williams maze were investigated. The moving belt task involved the rats walking

on a moving belt, where the surrounding area is electrified so that if a paw is placed off

the belt in any direction the animal will receive a shock. It was reported that tasks after

the 1-month washout phase, none of the 10mg/kg dosing regimes produced any

significant effect in any of the in comparison to the control but the 20mg/kg group

performed significantly slower at the Hebb-Williams maze and spent significantly more

time off the moving belt than the controls.

Stiglick and Kalant (1982) examining the effects of chronic cannabinoid

administration on the performance in an 8 or 12-arm radial maze. Rats were dosed orally

with an ethanolic cannabis extract so that they received 20mg/kg daily for 3 or 6 months.



After a 1-month washout phase, the animals that were dosed for 3-month were tested

with an 8 or 12-arm radial maze, and the animals that were dosed for 6-months were

tested with an 8-arm radial maze. It was reported that after the 1-month washout phase,

all cannabinoid treated animals learned the radial arm maze significantly slower than the

vehicle treated controls. It was reported that the control and cannabinoid treated animals

did not eat significantly different amounts and hence this result is unlikely to be because

of a lowered motivation to eat food caused by cannabinoid treatment.

Stiglick and Kalant (1983) conducted a similar experiment, although this time

pure THC was used and the behaviour of rats chronically treated with THC in a two-way

shuttle box avoidance test and a 12-arm radial maze was also investigated. Rats were

treated orally with 20mg/kg THC daily for 3 months. Testing in the 12-arm radial maze

began after a 34-day washout phase. Rats were then tested in an open field test after a 77-

day washout phase, then a differential reinforcement of low rate responding-20 (DRL-20)

test after a 92-day washout and finally the shuttle box avoidance test after a 132-day

washout. It was reported that treated with THC had no effect on open field test

exploratory behaviour in comparison to control. It was shown that THC treatment

effected the animals’ ability to learn the 12-arm radial maze in comparison to the vehicle

treated control, but that this effect was restricted to the first 11 days of performance. It

was reported that THC treated animals received significantly less food rewards during the

DRL-20 test in comparison to controls, but that this effect only last for the first 13 days of

testing. It was shown that THC treatment had no significant effect on the performance in

the shuttle box avoidance task.



Nakamura et al ., (1991) investigated the effects that chronic THC administration

had on performance in a variation of the 8-arm radial maze in the rat. Rats were dosed

with 5mg/kg THC I.P. 6 days a week, for 90 days. During administration the rats were

trained in an 8-arm radial maze where, after they had entered 4 arms, the rats were

removed from the maze for 5 seconds or 1 hour, and the placed back in the center. During

treatment, THC treated animals made significantly more errors in both the 5-second and

1-hour delay than the vehicle treated controls. 15 days after discontinuation of THC

treatment the THC treated animals still made significantly more errors than the controls,

but after 30 days the difference was no longer significant. Because the behavioural deficit

corrected after such a short time, it could be possible that it was just due to residual THC.

Unfortunately, plasma THC was not recorded, so this possibility can not be addressed.

Paule et al ., (1992) investigated the effect of cannabinoids on two operant tasks in rhesus

monkeys. Rhesus monkeys were exposed to the smoke of 1 2.6% cannabis cigarette 2 or

7 days a week, the smoke of 1 ethanol extracted cannabis cigarette 7 days a week or no

smoke for 1 year. Before exposure, animals were then trained approximately 30 times on

two tasks, a progressive ration (PR) operant test, where the number of presses to receive

the reward increased after each reward, but reverted back to 1 press at the end of each

day. The other task was a Conditioned Position Responding (CPR) test, involving

pressing a button in response to a light to receive a food reward where the correct button

changed depending on the color of the light. During dosing, the animals either continued

training (the active group) or were not trained (the residual group). In the active group, by

the end of their 12 months of treatment, the cannabinoid-exposed animals received

significantly fewer rewards or performed significantly slower that the cannabinoid free



animals in the PR, and CPR experiments. After the cannabinoid exposure the animals

were studied for 7 months where they received no drugs, during this time, the residual

group began training again. The data for the active group was not fully reported, but the

residual group still showed significantly impaired in performance in the CPR task after a

7 month abstinence period.

Human Neurological Imaging

Campbell et al., (1971) investigated the possibility of cannabis-induced cerebral

atrophy using pneumoencephalograms, a technique where the cerebral ventricles are

inflated with air in order to display their size. They found that subjects who had used

cannabis ‘consistently’ showed markedly bigger ventricles than the controls, and hence

cerebral atrophy. This study was seriously flawed as the cannabis using subjects were

polydrug abusers, which the controls were not, they were all under psychiatric treatment,

which the controls were not and worst of all, the first 4 cannabis using subjects were

know to have abnormal scans at the start of the trial. With the invention of the CAT scan

in the late 1970s, two studies used this far more sensitive technique to investigate the

claims of Campbell et al ., (1971).

Co et al ., (1977) and Kuehnle et al ., (1977) used CAT scans to look for any gross

cerebral atrophy in 12 and 19 chronic cannabis users respectively. Despite a lack of

control for any factor apart from sex, there were found to be no significant changes in

cerebral volume in comparison to controls.

Block et al ., (2000a) were the first to investigate the possibility of brain atrophy

induced by chronic cannabis use using magnetic resonance imaging. The cannabis using



subjects had used cannabis for at least 2 years, on average every day. It was found that

the cannabis users had no significant differences in gross brain, frontal lobe, temporal

lobe, parietal lobe, occipital lobe, cerebellar and hippocampal volume in comparison to

the controls.

Block et al ., (2000b) investigated the cerebral blood flow of abstinent chronic

cannabis users, using positron emission tomography (PET). The same cohort as Block et

al ., (2000a) received supervised abstinence from cannabis, alcohol and caffeine use for

an average of 44 hours, and a minimum of 33 hours before the PET scan. It was shown

that cannabis users had significantly lower regional blood flow in the posterior

cerebellum than the controls. Although this could be attributed to residual drug levels,

this seems unlikely as an experiment by the same laboratory reported that the cerebellar

blood flow increased during acute THC treatment (O'Leary et al., 2002). The possibility

that this decrease in blood flow is an effect of withdrawal can not be excluded until the

experiment is repeated with a longer period of abstinence.

Human Neuropsychological

There is a large history of neuropsychological testing in chronic cannabis users,

and a review the entire body of research is outside the scope of this article (for a review

see Gonzalez et al., 2002) so the most recent and illustrative examples are cited here.

Pope and Yurgelun-Todd (1996) compared the cognitive function in heavy and

light cannabis using college students. The rational behind the study was that if heavy

users were compared to light users then the difficulty of matching controls would be

avoided, as it was presumed that light and heavy cannabis users would come from the



same socio-economic background. Light users were defined as subjects who had smoked

cannabis a maximum of 9 times in the last month, and had no urinary traces of

cannabinoids. Heavy users were defined as subjects who had smoked cannabis a

minimum of 22 times in the last month and had traces of cannabinoids in their urine.

Subjects had 19 hours of supervised abstinence before they received a battery of

cognitive tests. It was reported that the heavy users did not performed significantly worse

than the light users in any test apart from the Wisconsin card sorting test and the

California verbal learning test. Interestingly, these results did not reach significance if the

genders were compared individually. Because of the short abstinence period, this

difference could be attributed to both residual cannabinoid action (because if there were

cannabinoids in the urine, there could be cannabinoids in the blood) or withdrawal

effects.

Fletcher et al ., (1996) investigated the effects chronic cannabis use has on

cognitive function in a young and an older group of Costa Rican men. The older group

had a mean age of 45 years old and had smoked for an average of 34 years. The young

group had a mean age of 28 and had smoked for an average of 8 years. The controls were

appropriately matched in regards to age, sex and socio-economic background. The

subjects were required to abstain from cannabis use for 72 hours, and any subjects who

had cannabinoids in their urine were excluded. Short-term memory, working memory,

and attentional skills were measured in each subject. It was reported that the older group

showed a significant deficit in short-term memory in comparison to the control group, but

no difference of any kind was seen in the young group. It seems unlikely that this result is

because of residual cannabinoids, as the urine tests were negative for cannabinoids. It



also seems unlikely that this effect was caused by cannabinoid withdrawal as withdrawal

symptoms last for 7 days after the end of cannabinoid administration (Smith, 2002) but

cannabinoids persist in urine for longer than this especially in heavy users (Ellis et al.,

1985). Therefore, if subjects had negative urine samples, they must have stopped

cannabis use a long enough time ago, that withdrawal symptoms would have passed.

Pope et al ., (2001) investigated the cognitive function of chronic cannabis users

over a 28-day abstinence period. There were 3 groups: subjects who had used cannabis at

least 5000 times and were smoking daily at the start of the study, subjects who had used

cannabis at least 5000 times but had used cannabis 12 times or less in the last three

months and the control group, who had used cannabis no more than 50 times. The

subjects were tested on a battery of tests on day 0, 1, 7 and 28 of abstinence. The former

users showed no significant difference in any of the tests. The current users showed

significant differences in many of the tests on day 0, 1 and 7, but by day 28 there was

only one test where they performed significantly worse, but when the test was controlled

for verbal IQ, the difference was no longer significant. The experiment shows that

washout phases of 19-72 hours, as used in other experiments probably are not long

enough to remove the residual effects of cannabinoids. Interestingly, the mean time of

use for the cannabis using group was 13 years. When compared to the results of Fletcher

et al ., (1996) this time length is far closer to the young group (9 years) who showed no

negative cognitive impact from cannabis use. Perhaps this indicates that cannabis use

needs to continue for more than 13 years to be damaging.



Summary

The results from the various experiments investigating chronic cannabinoid

treatment on neuronal morphology are summed up in table 1. In the rat experiments

reported here, all but one reported that cannabinoids alters neuronal morphology,

particularly in the hippocampus, and the one study that did not, used an unknown sample

size and techniques. In the rhesus monkey, the results have been inconclusive, but seem

to show that 1 2.6% THC cannabis cigarette a day is not enough to induced

neurohistological changes.

Studies investigating the neurochemical effect of chronic cannabinoid treatment

have had mixed results (Table 2). It has been reported that administration of cannabinoids

to pregnant rats produced neurochemical alterations in their offspring up to 40 days after

weaning (Walters & Carr, 1986), but when this experiment was repeated the alterations

found were completely different (Walters & Carr, 1988). Other experiments have found

no significant alterations in the neurochemical systems investigated in either the rat or the

rhesus monkey. It is possible that there is a critical period in which cannabinoids can alter

the neurochemistry of the rat.



T a b l e 1 . S t u d i e s i n v e s t i g a t i n g t h e e f f e c t s o f c a n n a b i n o i d s o n n e u r o

n a l m o r p h o l o g y

C h a n g e ? +

- - + + - +

T a b l e 2 . S t u d i e s i n v e s t i g a t i n g t h e e f f e c t s o f c a n n a b i n o i d s o n

n e u r o c h e m i s t r y

C h a n g e ? + + - - -

W a s h o u t P h a s e

8 m o n t h s

7 m o n t h s

N o n e

7 m o n t h s

N o n e

N o n e

n o n e

W a s h o u t P h a s e

0 - 4 0

d a y s

0 - 4 0

d a y s

0 - 2 w

e e k s

2 h o u r s - 2 m o n t h s

7 m o n t h s

6 0

d a y s

7 m o n t h s

D o s i n g L e n g t h

3 m o n t h s

1 y e a r s

9 0 d a y s

9 0 d a y s

8 m o n t h s

1 3 o r 2 4 m o n t h s

2 1 d a y s


~ 6 0 d a y s

~ 6 0 d a y s

9 0 d a y s

9 0 d a y s

1 y e a r

9 0 d a y s

1 y e a r

D o s i n g r e g i m e

1 - 3 2 . 6 % T H C c a n n a b i s c i g a

r e t t e s o r 0 . 7 m g / k g T H C I . V . d a i l y

1 2 . 6 % T H C c a n n a b i s c i g a r e t t e d a i l y

0 . 1 - 1 m g / k g T H C I . V . d a i l y

2 0 - 6 0 m g / k g T H C o r a l l y 5 x w e e k

4 - 8 m g / k g T H C o r a l l y 5 x w e e k

0 - 5 0 0 m g / k g T H C o r a l l y 5 x w e e k

2 m g / k g W I N 5 5 , 2 1 2 - 2 S . C . 2 t i m e s d a i l y


2 0 m g / k g T H C o r a l l y d a i l y t o d a m s

1 0 m g / k g T H C a n d C B D , 1 m

g / k g Δ 8 - T H C o r a l l y d a i l y t o d a m s



1 2 . 6 % T H C c a n n a b i s c i g a r e t t e 2 - 7 x w e e k


1 2 . 6 % T H C c a n n a b i s c i g a r e t t e d a i l y

A n i m a l

R h e s u s M o n k e y


R h e s u s M o n k e y R a t R a t R a t R a t

A n i m a l R a t R a t

R a t R a t


R a t




A u t h o r s

H a r p e r e t a l . , 1 9 7 7

S c a l l e t e t a l . , 1 9 9 0

A

n d r e w s e t a l . , 1 9 8 9

S c a l l e t e t a l . , 1 9 8 7

L a n d f i e l d e t a l . , 1 9 8 8

C h a n e t a l . , 1 9 9 6

L

a w s t o n e t a l . , 2 0 0 0

A u t h o r s

W a l t e r s a n d C a r r , 1 9 8 6

W a l t e r s a n d C a r r , 1 9 8 8

A l i e t a l . , 1 9 8 9

A l i e t a l . , 1 9 9 1

W

e s t l a k e e t a l . , 1 9 9 1

T a b l e 3 . S t u d i e s i n v e s t i g a t i n g

t h e e f f e c t s o f c a n n a b i n o i d s o n a n i m a l b e h a v i o u r

C h a n g e ? + + + ? +

T a b l e 4 . S t u d i e s i n v e s t i g a t i n g t h e e f f e c t s o f c a n n a b i n o i d s o n n e u r o l o g i c a l i m

a g i n g i n h u m a n s

C h a n g e ? + - - - +

T a b l e 5 . S t u d i e s i n v e s t i g a t i n g t h e e f f e c t s o f c a n n a

b i n o i d s o n n e u r o p s y c h o l o g i c a l t e s t i n g i n h u m a n s

C h a n g e ?

- - ?

W a s h o

u t P h a s e

1 m o n t h

1 m o n t h

7 7 - 1 3 2 d a y s

1 5 - 3 0 d a y s

7

m o n t h s

W a s h o

u t P h a s e

N o n e

N o n e

N o n e

N o n e

4 4 h o u r s

W a s h o

u t P h a s e

1 9 h o u r s

7 2 h o u r s

0 - 2 8 d a y s


3 0 - 1 8 0 d a y s

3 - 6 m o n t h s

3 m o n t h s

9 0 d a y s

1 y e a r

I m g a i n e T e c h n e q u e

p n e u m o e n c e p h a l o g r a m

C A T

C A T

M R I

P E T

H i s t o r y o f c a n n a b i s u s e

L i g h t u s e r s : U s e d < 9 t i m e s i n l a s t m o n t h . H e a v y u s e r s : u s e d > 2 2 t i m e s l a s t m o n t h

O l d u s e r s : U s e d f o r 3 4 y e a r s . Y o u

n g u s e r s : U s e d f o r 9 y e a r s

E x - u s e r s : U s e d > 5 0 0 0 t i m e s , b u t n o t i n l a s t 3 m o n t h s . C u r r e n t u s e r s : U s e d > 5 0 0 0 t i m e s


1 0 - 2 0 m g / k g T H C d a i l y o r a

l l y

2 0 m g / k g T H C d a i l y o r a

l l y

2 0 m g / k g T H C d a i l y o r a

l l y

5 m g / k g T H C 6 x w e e k I . P .

1 2 . 6 % T H C c i g a r e t t e 2 - 7 x w e e k

H i s t o r y o f c a n n a b i s u s e

“ C o n s i s t e n t ” u s e

“ C h r o n i c ” u s e

“ C h r o n i c ” u s e

D a i l y u s e f o r 2 y e a r s

D a i l y u s e f o r 2 y e a r s

A n i m a l

R a t

R a t R a t R a t

R h e s u s M o n k e y s



A u t h o r s

F e h r e t a l . , 1 9 7 6

S t i g l i c k a n d K a l a n t , 1 9 8 2

S t i g l i c k a n d K a l a n t , 1 9 8 3

N a

k a m u r a e t a l . , 1 9 9 1

P a u l e e t a l . , 1 9 9 2

A u t h o r s

C a m p b e l l e t a l . , 1 9 7 1

C o e t a l . , 1 9 7 7

K

u e h u l e e t a l . , 1 9 7 7

B l o c k e t a l . , 2 0 0 0 a

B l o c k e t a l . , 2 0 0 0 b

A u t h o r s

P o p e a n d Y

u r g e l u n e t a l . , 1 9 9 6

F l e t c h e r e t a l . , 1 9 9 6

P o p e e t a l . , 2 0 0 1

Experiments investigating the behaviour of rats and rhesus monkeys have

reported that it is altered, even after a significant washout phase, after chronic

cannabinoid treatment (Table 3). The one experiment which did not show this

conclusively was Nakamura et al ., (1991), which showed that the behavioural effects

were reversed after a 30 day washout, but not a 15 day washout phase. It is unknown

whether this is due to the persistence of cannabinoids in the plasma. Largely though, it

has been shown that cannabinoids in doses between 5 and 20mg/kg produce a residual

change in behaviour in the rat, and that this change is negative in nature, as it involves a

decrease in spatial processing as shown by reduced performance in 8 and 12-arm radial

mazes. In order to be conclusive about the action of chronic cannabinoids in monkeys,

more experiments need to be run.

In humans it has been shown that cannabinoids probably do not produce any

change in brain volume (Table 4). Block et al ., (2000b) reported that during abstinence,

cannabinoids produced a change in regional brain flow but the abstinent period was

probably not long enough to conclude that this was not an effect of residual cannabinoids

in the blood, or because of cannabinoid withdrawal.

Investigations into the cognitive state of cannabis users have shown very

interesting results (Table 5). The results of Pope and Yurgelun-Todd (1996) which



indicated that chronic cannabinoid usage resulted in cognitive impairment was probably

due to an insufficient washout phase. Fletcher et al ., (1996) reported that even after a

washout phase that was probably long enough for residual cannabinoids to be cleared and

withdrawal symptoms to subside, subjects who had used cannabinoids for a mean length

of 34 years performed significantly worse than their age matched controls in short term

memory tasks. On the other hand, it was reported that subjects who had used for a mean

of 9 years did not. This experiment indicates that perhaps significant cognitive

impairment only happens after extended cannabinoid treatment. Pope et al (2001) showed

that subjects who had used cannabis more than 5000 times but had been generally

abstinent for the last 3 months performed no worse than the controls. They also reported

that while subjects who had used cannabis more than 5000 times and were still using

daily performed significantly worse than the controls, they performed no worse than the

controls after 28 days abstinence.

As mentioned earlier, in order for a chemical to satisfy to ICON definition for

neurotoxicity, it must produced a physical change (neurochemical/neurohistological/

neurological) and this change must be associated with a negative behavioural effect.

Cannabinoids have satisfied this criterion in doses between 8 and 20mg/kg in the rat, but

not in the rhesus monkey. In humans there is no clear evidence that chronic cannabinoid

physical alters the brain or cognition, so there is no evidence that cannabinoids are

neurotoxic.

Even though cannabinoids have been shown to be protective against many

induced neurotoxic events (for review, see Fowler, 2003) in a model proposed by



Guzman (2003), it could be possible for cannabinoids to be protective against large

neurotoxic insults, yet cause neurotoxicity slowly over time.

In order to gain more knowledge in area, studies investigating rhesus monkeys

that are dosed in a more controlled fashion than smoke, and at a higher dose than 1

cannabis cigarette could prove useful. Instead of getting further away from human users,

this could in fact bring the experimental paradigm closer to human users, who often

smoke more than one cannabis cigarette a day (Foltin et al., 1989). Also studies

investigating the cognitive function of humans who have used cannabis for 30 years or

longer could show whether permanent cognitive impairments can be induced by extended

cannabis use.

In conclusion, there is evidence that chronic cannabinoid treatment is neurotoxic

to the rat, but not to the rhesus monkey or to humans.

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