Upregulation of mu opioid receptors by voluntary morphine administration in drinking water.

Morphine was provided to rats in drinking water for 21 days. Profound analgesic tolerance was detected both in hot-plate and tail-flick tests. The density of [3H]DAMGO binding sites increased by 76% in spinal cord membranes due to morphine exposure compared to those in opioid naive animals. Slightly augmented [3H]DAMGO binding was measured in the synaptic plasma membranes, with a concomitant decrease in the microsomal membranes, of morphine tolerant/dependent brains. These observations suggest that the regulation of spinal mu opioid receptors might be different from those in the brain. It is emphasized that the molecular changes underlying tolerance/dependence are influenced by several factors, such as the tissue or subcellular fractions used, besides the obvious importance of the route of drug administration. Results obtained after voluntary morphine intake further support the growing number of experimental data that chronic morphine does not internalize/downregulate the mu opioid receptors in the central nervous system.

Comparable system-level organization of Archaea and Eukaryotes.

A central and long-standing issue in evolutionary theory is the origin of the biological variation upon which natural selection acts. Some hypotheses suggest that evolutionary change represents an adaptation to the surrounding environment within the constraints of an organism's innate characteristics. Elucidation of the origin and evolutionary relationship of species has been complemented by nucleotide sequence and gene content analyses, with profound implications for recognizing life's major domains. Understanding of evolutionary relationships may be further expanded by comparing systemic higher-level organization among species. Here we employ multivariate analyses to evaluate the biochemical reaction pathways characterizing 43 species. Comparison of the information transfer pathways of Archaea and Eukaryotes indicates a close relationship between these domains. In addition, whereas eukaryotic metabolic enzymes are primarily of bacterial origin, the pathway-level organization of archaeal and eukaryotic metabolic networks is more closely related. Our analyses therefore suggest that during the symbiotic evolution of eukaryotes, incorporation of bacterial metabolic enzymes into the proto-archaeal proteome was constrained by the host's pre-existing metabolic architecture.

The large-scale organization of metabolic networks.

In a cell or microorganism, the processes that generate mass, energy, information transfer and cell-fate specification are seamlessly integrated through a complex network of cellular constituents and reactions. However, despite the key role of these networks in sustaining cellular functions, their large-scale structure is essentially unknown. Here we present a systematic comparative mathematical analysis of the metabolic networks of 43 organisms representing all three domains of life. We show that, despite significant variation in their individual constituents and pathways, these metabolic networks have the same topological scaling properties and show striking similarities to the inherent organization of complex non-biological systems. This may indicate that metabolic organization is not only identical for all living organisms, but also complies with the design principles of robust and error-tolerant scale-free networks, and may represent a common blueprint for the large-scale organization of interactions among all cellular constituents.

Ultrastructural localization of Hsp-72 examined with a new polyclonal antibody raised against the truncated variable domain of the heat shock protein.

In spite of the intensive search for the determination of the continuously widening physiological and pathological roles of different stress proteins, their ultrastructural localization at the electron microscopic (EM) level has hardly been examined. As it becomes increasingly evident that the function and physiological effectiveness of stress proteins are highly dependent on their spatial location and their associations with diverse regulator proteins, the demand for morphological studies which can identify their detailed distribution within the cells is evident. The reason for the practical lack of studies carried out at the EM level, lies in the shortage of reagents with suitable specificity and avidity necessary for this type of examination. To create such a reagent, a polyclonal antibody was raised using a recombinant truncated form of the inducible Hsp-72 protein. The antibody was extensively characterized, using different immunochemical methods to determine and verify its specificity, and then it was tried in ultrastructural examinations. Using the new antibody, it was possible to analyze the intracellular distribution of Hsp-72 with the immunogold technique. The localization of Hsp-72 was demonstrated directly at the ultrastructural level in the cytoplasm (especially at the cisterns of the RER), in the nucleus (mainly around the heterochromatic regions) and at both sides of the nuclear envelope close to the membrane pores. Apart from these localizations, Hsp-72 was found in several membrane bordered intracellular structures, which mainly belong to the endosomal-lysosomal system. We provide the first morphological verification of the appearance of Hsp-72 on the surface of the cells. Also novel is the indication, that the stress protein may recycle from the cell surface using a common route which includes coated pits and the endosomal system.