No abstract

Selective PCR amplification of human and mouse genomic DNAs with oligonucleotides encoding highly conserved regions of the S-opioid and somatostatin receptors generated a human DNA probe (hOPO1, 761 bp) and its murine counterpart (mOP86, 447 bp). hOPO1 was used to screen a cDNA library from human brainstem. A clone (named hORL1) was isolated, sequenced and found to encode a protein of 370 amino acids whose primary structure displays the seven putative membrane-spanning domains of a G protein-coupled membrane receptor. The hORL1 receptor is most closely related to opioid receptors not only on structural (sequence) but also on functional grounds: hORL1 is 49-50% identical to the murine ,u-, 6-and rc-opioid receptors and, in CHO-Kl cells stably transfected with a pRc/CMV:hORLl construct, ORLl mediates inhibition of adenylyl cyclase by etorphine, a 'universal' (nonselective) opiate agonist. Yet, hORL1 appears not to be a typical opioid receptor. Neither is it a somatostatin or o (N-allylnormetazocine) receptor. mRNAs hybridizing with synthetic oligonucleotides complementary to mOP86 are present in many regions of the mouse brain and spinal cord, particularly in limbic (amygdala, hippocampus, septum, habenula, . . .) and hypothalamic structures. We conclude that the hORL1 receptor is a new member of the opioid receptor family with a potential role in modulating a number of brain functions, including instinctive behaviours and emotions.

Abstract. The amyloid precursor protein (APP) is a transmembrane protein expressed in several cell types. In the nervous system, APP is expressed by glial and neuronal cells, and several lines of evidence suggest that it plays a role in normal and pathological phenomena. To address the question of the actual function of APP in normal developing neurons, we undertook a study aimed at blocking APP expression using antisense oligonucleotides. Oligonucleotide internalization was achieved by linking them to a vector peptide that translocates through biological membranes. This original technique, which is very efficient and gives direct access to the cell cytosol and nucleus, allowed us to work with extracellular oligonucleotide concentrations between 40 and 200 nM. Internalization of antisense oligonucleotides overlapping the origin of translation resulted in a marked but transient decrease in APP neosynthesis that was not observed with the vector peptide alone, or with sense oligonucleotides. Although transient, the decrease in APP neosynthesis was sufficient to provoke a distinct decrease in axon and dendrite outgrowth by embryonic cortical neurons developing in vitro. The latter decrease was not accompanied by changes in the spreading of the cell bodies. A single exposure to coupled antisense oligonucleotides at the onset of the culture was sufficient to produce significant morphological effects 6, 18, and 24 h later, but by 42 h, there were no remaining significant morphologic changes. This report thus demonstrates that amyloid precursor protein plays an important function in the morphological differentiation of cortical neurons in primary culture.

We suggest that a lack of nitric oxide synthase in pyloric tissue is responsible for pylorospasm in infantile hypertrophic pyloric stenosis.

Pleiotrophin (PTN), also known as HB-GAM, belongs to an emerging cytokine family unrelated to other growth factors. We report here the first comprehensive study using in situ hybridization on the cellular distribution of this new heparin-binding growth factor mRNA in rat tissues. PTN mRNA was developmentally expressed in many--but not all--neuroectodermal and mesodermal lineages, whilst no PTN mRNA was detected in endoderm, ectoderm and trophoblast. PTN mRNA was found in the nervous system throughout development, with a post-natal peak of expression. In the adult nervous system, significant expression persisted in hippocampal CA1 pyramidal neurons and in cortical neurons, but also in different non-neuronal cells types in various locations (olfactory nerve, cerebellar astrocytes, pituicytes, Schwann cells surrounding the neurons in sensory ganglia). PTN mRNA was also found during development in the mesenchyme of lung, gut, kidney and reproductive tract, in bone and cartilage progenitors, in dental pulp, in myoblasts, and in several other sites. Expression was differently regulated in each location, but usually faded around birth. In the adult, PTN mRNA was still present in the meninges, the iris, the Leydig cells of the testis and in the uterus. PTN mRNA was also strongly expressed in the basal layers of the tongue epithelium, which is the only epithelium and ectodermal derivative to express PTN mRNA, and this only after birth. PTN is known to be a growth factor for perinatal brain neurons and a mitogen for fibroblasts in vitro. Recently, trophic effects on epithelial cells and a role as a tumour growth factor have been reported. The mechanisms of regulation and the functions of PTN are however still uncertain. Its expression pattern during development suggests important roles in growth and differentiation. Moreover, the presence of PTN mRNA in several adult tissues and the up-regulation of PTN mRNA expression in the gravid uterus indicate that PTN also has physiological functions during adulthood.