James J. Thompson

FOS expression induced by interleukin-1 or acute morphine treatment in the rat hypothalamus is attenuated by chronic exposure to morphine

Interleukin-1 (IL-1) is a cytokine involved in a variety of biological activities. It has been hypothesized that the immunomodulatory effects of IL-1 are the result of both direct action on immune cells and indirect action on a regulatory cascade mediated through the hypothalamus. Chronic exposure to substances of abuse, such as morphine, appears to modulate immunoresponsiveness by mechanisms not yet defined. The expression of FOS, the protein product of the c-fos proto-oncogene, has been widely used as an anatomical marker for monitoring neuronal activity. We have previously shown that acute treatment with either morphine or IL-1 induces FOS immunoreactivity in the rat brain, including the paraventricular (PVN) and supraoptic (SON) nuclei of the hypothalamus. In this study, using immunocytochemical staining of FOS, we demonstrate that chronic exposure to morphine attenuates the cellular responsiveness to IL-1 and to morphine in the PVN and SON, whereas pretreatment with naloxone, an opiate receptor antagonist, does not reverse the effect of IL-1 on FOS expression. The results not only confirm that the PVN and SON are neuroanatomical sites where the actions of both morphine and IL-1 converge, but also indicate that chronic exposure to morphine may desensitize the cellular response involved in hypothalamic functions through an IL-1-dependent pathway.

Gradient acrylamide/agarose gels for electrophoretic separation of intact human very low density lipoproteins, intermediate density lipoproteins, lipoprotein a, and low density lipoproteins

An exponential gradient gel with 0-10% acrylamide and 0.5% agarose was developed for electrophoresis of intact high molecular weight lipoproteins. This system resolves very low density lipoproteins, intermediate density lipoproteins, lipoprotein a, and low density lipoproteins in a size-dependent fashion. The characteristic relative mobility of these species can be determined in relation to protein and colloidal gold reference materials. Electron microscopy of selected lipoprotein fractions confirmed that relative mobility was related to apparent lipoprotein diameter. The composite gel medium can be used with prestained lipoproteins and permits immunoelectroblotting for qualitative analysis of apolipoprotein constituents.

Morphine modulates IL-1 activation of fos in the rat hypothalamus

Abstract By using FOS proto-oncogene protein as a marker for neuronal activity induced by an extracellular stimulus, we have shown that several hypothalamic nuclei are sites of action for interleukin-1 (IL-1). These nuclei, including the paraventricular (PVN) and supraoptic (SON) nuclei, have relatively few IL-1 receptors. One of the possibilities for this dysjunction may be that IL-1 acts on receptors other than currently defined IL-1 receptors to induce FOS activation. Acute subcutaneous morphine treatment also increased FOS immunoreactivity in several hypothalamic nuclei including PVN and SON. Taken together, these results suggest the possibility of an interaction between morphine and IL-1 in the hypothalamic PVN and SON. The PVN and SON have been suggested to be involved in a neuroendocrine-immune axis. In this study we examined the effects of chronic morphine on IL-1 activation of FOS. The chronic morphine pretreatment consisted of 2 pellets on the 1st day and 4 pellets on the 2nd day, 75 mg morphine sulfate/pellet. This regimen has been shown to induce a high degree of tolerance and dependence, and it markedly decreased IL-1 activation of c-fos mRNA in the hypothalamus and FOS immunoreactivity in the hypothalamic PVN and SON nuclei. These data suggest that attenuation of IL-1 activation of FOS expression in the hypothalamus may be a mechanism by which morphine tolerance and dependence affects the neuro-endocrine-immune axis.

Efficient electroblotting of human very-low-density lipoproteins and low-density lipoproteins from composite gradient gels to nylon membranes

Rinsing of gradient composite acrylamide/agarose gels with 1% Triton X-100 permits the efficient electrotransfer of very-low-density and low-density lipoproteins from the gels and does not appear to interfere with subsequent capture of the lipoproteins by charged nylon membranes. Overall efficiency of the transfer/capture process can approach 95% and does not appear to be significantly affected by total lipoprotein concentrations up to 5000 mg/dl. Direct immunoquantification of transferred apolipoproteins on the membrane is feasible as well. The nylon membranes used, however, must be pretested to ensure capture efficiency.