A Study of Some of the Effects of Aqueous Leaf Extract of Cannabis sativa on the Visual Cortex of Adult Wistar Rats

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Introduction

Hallucination, in the broadest sense, is the subjective perception of an object or event when no such stimulus or situation is present1,2 . In a stricter sense, it is defined as a perception in a conscious and awake state in the absence of external Stimuli, which have qualities of real perception located in external objective space2. It can occur in any sensory modality: visual, auditory, olfactory, gustatory, tactile, proprioceptive, equilibrioceptive, nociceptive, thermoceptive and chronoceptive3. A mild form of it is called disturbance, and can occur in any of the senses. Hallucination may be benevolent (i.e. telling someone good things about himself or malicious(i.e. cursing someone) and occurs in both healthy and sick individuals4. The most common modality of hallucination is visual which includes the phenomena of seeing things which are not present. Different causes of visual hallucination have been classified as psychobiochemical, (a disturbance of neurotransmitters), psychophysiological (a disturbance of brain structure) and psychological/psychodynamic (e.g. meaningful experience intruding into consciousness5. Manford and Andermann (1998)6 summarized three pathophysiological mechanisms of visual hallucination as thus:

i. irritation of the visual cortex, both the primary (Brodmann's Area 17) and association (Brodmann's Areas 18 and 19)

ii. deafferentation of visual system which may lead to cortical release phenomenon and

iii. effects on reticular activating system

A lesion of any nature and from any source on the visual system, especially involving the visual cortex will result in visual hallucination. Studies have shown that drugs and plant extracts have association with visual hallucination. One of the important plants which have been attributed with visual hallucination is Cannabis sativa 7 .

Cannabis sativa, commonly called Marijuana or Indian Hemp is an annual plant in the Cannabaceae family and a herb that has been used throughout recorded history by Man as a source of fibres, for its oil, as food, as a drug and medicine and for spiritual purposes8 . It has its action on the higher nerve centres and can produce an exhilarating intoxication with hallucination7 . Its psychoactive property made it a widely used street drug in Nigeria despite the legal implication of its possession and use. The World Drug Report 2006 (WDR 2006) published by the United Nations Office on Drug and Crime 9 showed that the annual prevalence of cannabis use in Nigeria is about 13.8% as at year 20002 .It is to be noted that Annual Prevalence of Cannabis use is the percentage of youth and adult population (aged 15-64 years) who have consumed the drug once in the past survey year. Nigeria with this value, has the 9th highest prevalence of cannabis use in the whole world9 .

Methodology

600g of dried leaves of Cannabis sativa, obtained from the Kwara State Command of NDLEA was grinded with local mortar and pestle into powdery form. 100g of the power was soaked in 100ml of distilled water for 72hours and filtered after 72 hours with Whatman's No 1 filter paper to get 800mls of filtrate. The filtrate, was oven-dried at a temperature of 600c for 7 days to form a deep brown paste of 10g which was dissolved in 50mls of phosphate buffered saline to make a 200mg/ml aqueous solution of cannabis sativa.

Sixteen (16) adult Wistar Rats with average weight of 200g (weight range being 170g to 240g) were procured from the animal house of the Department of Zoology, University of Ilorin. They were of the two sexes with 8 rats being male and 8 female rats. Standard rat diet was purchased from Bendel Feeds, Taiwo Road, Ilorin.

The animals were reared in the animal holdings of the Faculty of Basic Medical Sciences of University of Ilorin. They were made to acclimatize for two (2) weeks before the commencement of administration of plant extracts. They were fed with standard rat diet which was purchased from Bendels feeds at once to avoid change in diet composition and also given tap water ad-libitum. They were given free assess to food and water. They were kept in standard laboratory wooden cages in groups of four (4) with male and female animals kept separately. They were generally cared for under standard laboratory conditions of good lighting, moderate temperature and adequate ventilation, and in a neat environment. They were weighed routinely everyday using Saltul Weighing balance.

The animals were divided into two (2) experimental and control groups A and B, each group of animals had four male and four female rats which were kept in separate apartments to prevent mating and pregnancy by the female animals. Animals in group A were treated with 300 mg/kg body weight/day (0.3ml) of Cannabis sativa for 21 days while group B animals were treated with phosphate buffered saline equal volume of the dose of each extracts (i.e. 0.3 ml) daily for 21 days. Administration of the extract and phosphate buffered saline was done orally with the aid of oro-gastric feeding tube at 07.00 hour each day.

The animals were sacrificed by cervical dislocation 24 hours after the 21st day of administration. Brain tissue was carefully removed from the skull and occipital cortex quickly excised for tissue homogenate preparation while the whole brain tissue was fixed in 10% formol calcium for 72 hours, after which right occipital cortexes were excised separately for further histological processing. The excised left occipital cortex was put in mortar (Lao Style) after being weighed with sensitive weighing balance. 1ml of 0.25M (6.625%) sucrose solution was added and the tissues homogenized thoroughly well. Tissue homogenate was collected in 5ml Plain serum bottle for enzyme assay.

The data was analysed using the computerized statistical package SPSS version 17. Mean and Standard Error of Mean (SEM) values for each group was determined. The Mean values of experimental and control groups were compared using student's t-test at 95% confidence interval to determine the level of statistical significance between the mean values

Results

The animals were closely observed throughout the period of the investigation. During acclimatization, all animals appeared presumably normal with smoothly laid hairs on their back and pinkish eyes. They fed well throughout. From the first day of administration of the plant extract, animals in the experimental group appeared restless for a few hours after administration. They had hairs on their back standing out with pinkish and radiating eyes. They appeared dull and inactive as the day passed by and withdrew from feed and water. They appeared presumably normal in the last 12hours of the day averagely and fed more. On approaching the end of administration, the animals in the experimental groups, especially appeared more docile and less active. The Group B animals (control group animals) appeared to maintain the pre-administration observation status throughout the period of this investigation.

Weights of the animals were monitored at 07.00 hour of every day and recorded. Before administration of the plant extract, all animals in both the experimental and control groups were steadily gaining weight. From the third day of administration, animals in the experimental group were observed to begin to lose weight. Their weights were stable the first and second day of administration (not increasing nor decreasing). Animals in the control group, however continued to have a steady weight gain. Statistical analysis of the weight changes indicated that there is no statistically significant difference in the weight changes of the animals in the experimental group, relative to those in the control group, as P >0.05. The average weight on selected days, mean, standard error of mean (SEM) and the level of significance at 95% confidence interval are represented on tables 1, 2 and 3.

There was an increase in the homogenate level of LDH in the experimental animal, compared to control animals, with a statistically significant difference in the average level of LDH (P

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