Abstract

Introduction

Materials & Methods

Results

Discussion & Conclusion

References

Discussion
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While the mean basal DBT was not altered with age, the data here indicate that mean basal metabolic rate (BMR) of young mice was significantly higher than the old mice of both subgroups. This indicates that there is an age-related decrease in BMR and oxygen consumption. Such a decrease is attributable to a decrease in fat-free mass and brown adipose tissue with age (Horan et al, 1988; Pratley et al, 1994; Toth et al, 1994). A comparison of mean BMR in the two subgroups of old mice indicated that there was no statistically significant difference between these two subgroups of old mice. Therefore, although basal DBT of old mice was similar to that of the young mice, the BMR of old mice was significantly less than young mice. This suggests that old animals had the capability of maintaining DBT under normal circumstances even though their BMR was reduced compared to young animals.

It has been shown previously that oxygen consumption rises as the ambient temperature is dropped from 20 °C down towards zero (Ganong, 1987). Similar results were obtained with all animal groups where oxygen consumption initially rose before exhibiting a decline. The young animals were able to maintain their MR showing a13% decline. Those old animals that were able to maintain their DBT like the young animals also exhibited a marginal drop in their oxygen consumption of 16%. A significant drop (80%) in oxygen consumption was noted in the group of old animals that were unable to maintin their DBT. Hence, clear differences were observed between old and young animals that accentuated with the cold challenge.

Age alone could not account for the differences observed in the cold challenge; one subgroup of old animals behaved like the young animals while the other did not. Therefore, it must be concluded that some biological mechanism other than the difference in MR must account for the differences observed. Since thermoregulation is centrally controlled, it was considered that a difference between the subgroups might be observed in hypothalamus and perhaps the hippocampus, as investigators previously had suggested an inter-relationship between hippocampus and hypothalamus for the effects of IL-1 (Hopkins and Rothwell, 1995; Rothwell, 1991; Blatteis, 1992).

IL-1 causes alteration of the thermosensitive neurons by increasing the firing rate of the cold-sensitive neurons and depressing the firing rate of the warm-sensitive neurons of the anterior hypothalamus and preoptic area, and thus resetting the thermostat at an elevated level (Hori et al, 1988). This evidence together with the known role of hypothalamus in thermoregulation strongly suggests that IL-1 may play a role in maintenance of DBT under such physiological conditions as ageing and thermal stress. Age related changes in expression and receptors have also been suggested by the finding that an increased IL-1 expression and IL-1 receptor density has been reported in such neurodegenerative conditions as Alzheimer’s disease and Down’s syndrome, which are generally associated with ageing (Giulian et al, 1985; Griffin et al, 1989; Cacabelos et al, 1994).

Most of the data available on IL-1 receptors is based on recombinant IL-1a because of the poor binding capacity of [125I] IL-1ß and higher ligand concentration required (Ban et al, 1991). [125I] IL-1ß was used in the present experiments because the pyrogenic effects of IL-1 are shown to be blocked by an antibody to the Type II receptor (Luheshi et al, 1993), and it is known that among the two types, IL-1ß has been shown to bind preferentially to Type II receptors (Scapagliata et al, 1989).

The data reported here indicate that there was no specific binding of IL-1ß in hippocampus and hypothalamus of young animals. This observation is consistent with previous data as non specific binding of IL-1a in mouse hippocampus has been reported by two independent groups of investigators (Parnet et al, 1994 and Takao et al, 1990), and was considered to be nearly 60% of total binding (Takao et al, 1990). It has also been reported that specific binding for IL-1ß is less marked than that of IL-1a (Ban et al, 1991 and Haour et al, 1990), as low as 24 times (Takao et al, 1990).

Those old animals which were unable to maintain DBT above 18 °C in the cold challenge, showed the highest binding in both hippocampus and hypothalamus compared to the old and young animals of the other groups. In both brain areas there was evidence of specific binding. The appearance of specific binding following the cold challenge may be due to the fact that the stress in these animals led to a greater release of glucocorticoids, up-regulating IL-1 receptors and thus resulting in enhanced IL-1ß binding.

In a thermal challenge it is likely that besides release of glucocorticoids (McElwen et al, 1987), release of endogenous IL-1 also occurs, since it has been reported that expression and activity of cytokines, including IL-1, is increased in conditions of tissue stress (Shintani et al, 1995; Cooper and Rothwell, 1994; Hopkins and Rothwell, 1995). Glucocorticoids tend to up-regulate IL-1 receptors and decrease production of IL-1, but the direct increase in the endogenous IL-1 production secondary to the thermal challenge in turn may cause a down-regulation of IL-1 receptors (Haour et al, 1992). Glucocorticoids released via an action of IL-1 through CRF and ACTH (Tsagaraki et al, 1989; Katsuura et al, 1990), are likely to result in negative feedback on further release of glucocorticoids at multiple levels (Rothwell, 1991; Rothwell, 1992 and Cooper and Rothwell, 1994). This intertwined role of IL-1 and glucocorticoids is probably designed to limit increase in the receptor numbers. Despite this control, the evidence indicates that an increased hypothalamic and hippocampal binding occurred in the cold challenged animals. It must be noted that the changes failed to reach statistical significance when an alpha level of 0.05 was chosen but “p” values between 0.10 and 0.05 were observed in most of these cases.

When the data obtained from all animals was combined to investigate the effect of the cold challenge alone, it was evident that there was a marked increase in IL-1ß binding in both hippocampus and hypothalamus. Specific binding became apparent indicating that hypothalamic and hippocampal IL-1ß receptors are up-regulated in stressful situations like a cold challenge. However these data may also be indicative of a role for IL-1ß in temperature regulation at the hypothalamic level and perhaps also at the level of hippocampus.

The present results obtained from immunohistochemical analysis are consistent with the literature in stating that IL-1 expression is present in hippocampus. One group of investigators have reported immunoreactive material in apical dendrites of pyramidal cells in the field of CA3 and CA4 and a particularly dense network of fibres around the pyramidal cells in CA1 region of the hippocampus (Lechan et al, 1990). Using in situ hybridization studies, Jirikowski and co-workers (1990) and Bandtlow and co-workers (1990), were able to show localisation of IL-1ß mRNA in the pyramidal cell layer of the hippocampus. This raised the possibility that if pyramidal cells do not show IL-1 expression in immunohistochemical analysis but IL-1ß mRNA can be detected, then there is either transcription of IL-1 DNA without translation to protein, or minimum translation under normal circumstances, limiting detection of IL-1 by immunohistochemistry. These experiments confirm that IL-1ß is expressed in pyramidal cells of hippocampus.

Expression of IL-1ß after subjecting animals to a cold challenge show that there was an increased IL-1ß expression in hypothalamus of both old and young animals when they are exposed to a cold challenge. As in the case of IL-1ß receptor binding, this may result from stress induced by the cold challenge. An alternative possibility is that expression is increased so that DBT is maintained to a reasonable level. In this case, it might be speculated that IL-1ß expression in hypothalamus plays a role in thermoregulation. A further increase in IL-1ß expression in hypothalamus of old compared to the young animals perhaps suggests an additional role in old animals. The fact that this expression is further enhanced in those old animals which were unable to maintain DBT above 18 °C in the cold challenge may support this hypothesis. Old animals may also be more susceptible to glucocorticoids and thus resulting in their inability to maintain DBT (Strijbos et al, 1993). These data parallel the data obtained from IL-1ß binding experiments, showing an increased density of these receptors in old animals and especially an increased binding in those old animals which were unable to maintain DBT above 18 °C in the cold challenge.

The reason that some old animals, although exhibiting increased IL-1ß binding and expression, failed to maintain DBT may be due to the fact that Type II receptor to which IL-1ß binds preferentially (Luheshi et al, 1993; Cunningham and DeSouza, 1993), has not only been shown to be more actively up-regulated by glucocorticoids (Colotta et al, 1993; Nicola, 1994), but is also susceptible to be shed (most likely through protein cleavage) by glucocorticoids (Colotta et al, 1993; Nicola, 1994). Therefore, IL-1ß bound to this receptor is rendered function-less. It has also been suggested by Colotta and co-workers (1993) that Type II receptor is a decoy receptor for IL-1ß, not able to produce any of the IL-1 induced responses, and thus traps IL-1ß in order to control IL-1ß generated responses through Type I receptor. However, this hypothesis is not shared by Luheshi and co-workers (1993) who have demonstrated Type II receptor to be responsible for fever and thermogenesis. Another possible post-receptor event responsible for the lack of increase in DBT in some old animals may be secondary to a diminished release of PGE2 from the brain (Grahn et al, 1987).

Conclusion.

Old and young mice behave differently under a thermal stress. Young animals maintain DBT better than the old animals. The old animals were subdivided on the basis of their ability to maintain DBT. Some behaved like young animals and were able to maintain DBT greater than 18 °C, while in other old animals DBT was further reduced. IL-1ß receptor density and IL-1ß expression are age-related and relate to the ability to sustain a cold challenge.

These experiments confirm the presence of receptors for IL-1ß in mouse hypothalamus and hippocampus. Receptor density is higher in hippocampus than hypothalamus. In addition IL-1ß expression was observed in hypothalamus and hippocampus. In hippocampus IL-1ß expression was marked in pyramidal cells compared to the dentate gyrus granule cells.

On the basis of the data presented here, it is suggested that old animals have a higher density of IL-1ß receptors and IL-1ß expression than the young animals. In response to the cold challenge receptor density and IL-1ß expression increase in young and old animals but the increase observed was more enhanced in old animals and especially those old animals which were unable to maintain DBT above 18 °C in the cold challenge.

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