Analgesic, learning and memory and anxiolytic effects of insulin in mice

Abstract

Insulin is a polypeptide hormone that is present in mammals and its main function is the maintenance of adequate blood sugar level. Insulin receptors are widely but unevenly distributed in the brain. Insulin has been reported to be involved in the regulation of neurotransmitters release. It has also been linked to the pathogenesis of neurodegenerative disorders such as Alzheimer's and Parkinson's diseases. Although there is abundant literature on the study of biochemical and molecular properties of insulin, there has been no literature on its central behavioural effects on anxiety and pain relief among other behavioural effects. This study therefore investigates whether insulin has any anxiolytic and other CNS effects.

This experiment was carried out in mice using animal behavioural models including a hot plate analgesic test, holeboard and elevated plus maze for anxiolytic test. A Y-maze was used for the locomotor activity and spontaneous alternation investigations. Mice were administered intraperitoneally with insulin at different doses of 0.5, 1.0 and 2.0 IU/kg.

The results obtained showed that insulin has no analgesic activity, however, it caused significant central inhibitory effects by decreasing both locomotor activity in both holeboard and Y-maze models and also decreased the exploratory behaviour in holeboard at doses administered dose-dependently indicating its sedative effects. In elevated plus maze, insulin had no effects on percentage of open arm entries at all doses but had a significant effect on percentage of open arm duration at the dose of 1.0 IU/kg only. Insulin administration at lower doses (0.5 and 1.0 IU/kg, i.p.) had no effect on spatial working memory, however, it had significant spatial working memory impairment at the dose of 2.0 IU/kg, i.p. in mice. The study showed that insulin has several neuropharmacological effects at doses used.

Introduction

Insulin is an acidic proteinous hormone having a molecular weight of approximately 5600. This hormone is composed of two polypeptides called A and B chains which are covalently joined by two intrachain disulfide bonds. A third intrachain disulfide bridge is present in the A chain. The chains are formed by proteolysis of pro-insulin, a larger single chain precursor, by removal of the intervening sequence of amino acids, referred to as the C-peptide. Insulin is essential for metabolic processes involving carbohydrates, fats and proteins. In most tissues, insulin stimulates the uptake of glucose, fatty acids and amino acids and aids their eventual conversion into storage forms. Naturally, higher animals produce their own endogenous insulin from the pancreas. Today there are various types of insulin, which are classed based on their onset of action, peak time and duration of action. Insulin is used in the management of diabetes mellitus, where there is either a lack of insulin production or non-sensitivity of the β-cells of the pancreas to produce insulin.

The existence of insulin receptors within the brain is less well known and the functions of these receptors are somewhat an enigma. It is important to note that brain cells are not fully reliant upon insulin for glucose supply, they have independent means of obtaining glucose. Also, brain insulin receptors (InsRb) differ somewhat from their peripheral counterparts [1]. The physiological role of insulin receptors in the brain appears to be twofolds: (1) the tight control of glucose transport in specific brain regions and (2) the yet incompletely understood function in the central nervous system (CNS) development and function. Aberrant function of the brain insulin receptors has been hypothesized to be involved in CNS dysfunction [29]. Even distribution of insulin receptors would be expected throughout the brain particularly if their only function is to mediate insulin-induced glucose transport into neurons as a source of energy. It has been shown that the highest brain-insulin receptor densities are found in the olfactory bulb, cerebral cortex, hypothalamus, cerebellum and choroids plexus [1], [2], [12], [16], [33]. Furthermore, high densities of brain insulin receptors are found in the thalamus, caudate putamen and some mesencephalic and brain stem nuclei. Insulin concentrations in extracts of whole brain averaged about 25-fold greater than the concentration in plasma [13], [14], [35].

The brain distribution of insulin receptor messenger RNA investigated using in situ hybridisation is consistent with the distribution of insulin receptors. Insulin brain receptor messenger RNA is most abundant in the cell layers of the olfactory bulb, in dentate gyrus of the pyriform cortex and hippocampus, in the choroid plexus and in the arcuate nucleus of the hypothalamus [1], [23].

Surprisingly, there is some mismatch between receptor density and insulin concentration. Hypothalamus and cerebral cortex have similar levels of receptor content but have more than a 4-fold difference in insulin content. Such disparity suggests that the insulin receptors may have other functions than those related to glucose utilization only [13], [14]. Recently, Hoyer et al. [17] demonstrated that the brain is sensitive to exogenous glucose metabolism because of the receptors in the brain. Furthermore, energy homeostasis, including control of food intake and energy expenditure, is affected by insulin levels [6], [24], [25]. Also, the presence of insulin in the brain has effects on behavioural functions such as food intake, motor activity and memory due to their classical role in metabolism [26]. These behavioural activities clearly support the hypothesis that insulin has central nervous system functions.

Insulin brain receptors and degenerative diseases have exhibited a certain relationship since the brain receptors are of higher densities on the hippocampus and parts of the cerebral cortex which are important in learning and memory in man. In disorders like Alzheimer's disease, Parkinsonism and Huntington's chorea, brain insulin concentrations, have been shown to decrease with age. It has been suggested that insulin should be administered in brain degenerative diseases to enhance memory in such patients [4], [7], [18], [22]. It appears that insulin has been implicated in energy deficits pertaining to these diseased states [3]. From the literature on the role of insulin in the brain, it is revealed that there still remains a large disparity in the knowledge about insulin possible cognitive effects.

Therefore, even though some studies have been carried out on the relationship between insulin and its effects on the brain [29], there are still areas of insulin-brain activity, which have not been studied. No direct studies of a possible relationship between insulin or insulin receptor with anxiety have been found in literature. Furthermore, insulin has been reported to be involved in energy homeostasis and deficiency in insulin can cause neuropathic pain [26] and changes in energy status in the brain could impact many behavioural effects. Thus, this experiment was designed to study the effects of insulin in the brain and to establish any possible relationship with pain, learning and memory and anxiety in mice using various behavioural animal models such as hot plate (central analgesic effect), Y-maze (spatial working memory effect), elevated plus maze and hole board (exploratory behaviour).