Significance of Autoantibodies in Autoimmune Encephalitis in Relation to Antigen Localization: An Outline of Frequently Reported Autoantibodies with a Non-Systematic Review.

Autoantibodies related to central nervous system (CNS) diseases propel research on paraneoplastic neurological syndrome (PNS). This syndrome develops autoantibodies in combination with certain neurological syndromes and cancers, such as anti-HuD antibodies in encephalomyelitis with small cell lung cancer and anti-Yo antibodies in cerebellar degeneration with gynecological cancer. These autoantibodies have roles in the diagnosis of neurological diseases and early detection of cancers that are usually occult. Most of these autoantibodies have no pathogenic roles in neuronal dysfunction directly. Instead, antigen-specific cytotoxic T lymphocytes are thought to have direct roles in neuronal damage. The recent discoveries of autoantibodies against neuronal synaptic receptors/channels produced in patients with autoimmune encephalomyelitis have highlighted insights into our understanding of the variable neurological symptoms in this disease. It has also improved our understanding of intractable epilepsy, atypical psychosis, and some demyelinating diseases that are ameliorated with immune therapies. The production and motility of these antibodies through the blood-brain barrier into the CNS remains unknown. Most of these recently identified autoantibodies bind to neuronal and glial cell surface synaptic receptors, potentially altering the synaptic signaling process. The clinical features differ among pathologies based on antibody targets. The investigation of these antibodies provides a deeper understanding of the background of neurological symptoms in addition to novel insights into their basic neuroscience.

Proliferative enteropathy (PE)-induced changes in the calbindin-immunoreactive (CB-IR) neurons of inferior mesenteric ganglion supplying the descending colon in the pig.

A subpopulation of the pig inferior mesenteric ganglia (IMG) neurons projecting to the colon exhibit calbindin-like immunoreactivity. It is not known if there are any changes in the chemical coding patterns of these neurons during porcine proliferative enteropathy (PE). To answer this question, juvenile Large White Polish pigs with clinically diagnosed Lawsonia intracellularis infection (PE; n = 3) and a group of uninfected controls (C; n = 3) were compared. The retrograde tracer fast blue (FB) was injected into the descending colons of all animals and then tissue comprising IMGs from both groups was processed for double-labeling immunofluorescence with calbindin-D28k (CB) in combination with either tyrosine hydroxylase (TH), neuropeptide Y (NPY), somatostatin (SOM), vasoactive intestinal polypeptide (VIP), nitric oxide synthase, Leu-enkephalin, substance P, vesicular acetylcholine transporter, galanin, or pituitary adenylate cyclase-activating polypeptide. Immunohistochemistry revealed changes in the chemical coding pattern of calbindin-immunoreactive neurons in the inferior mesenteric ganglia of the pig. In control animals, FB/CB-positive neurons were immunoreactive to TH, NPY, SOM, and VIP. In the experimental group, TH-expressing neurons were unaffected, NPY-expressing neurons were increased, whereas the number of neurons immunoreactive to SOM or VIP was reduced. Changes in chemical coding of CB neurons during PE may play an important role in adaptation of these IMG cells under pathological conditions.

The regulatory mechanism of neuromedin S on luteinizing hormone in pigs.

Neuromedin S (NMS) has been implicated in the regulation of luteinizing hormone (LH) secretion. However, the regulatory mechanism of NMS on LH in pigs remains unexplored. In the present study, we confirmed the hypothesis that the effect of NMS on LH could be mediated via hypothalamic melanocyte-stimulating hormones (MSH) neurons of ovariectomized pigs. In an immunohistological experiment, NMS receptor NMU2R-positive neurons were found in the paraventricular nucleus of hypothalamus, widely distributed in the anterior pituitary, and sparsely observed in the posterior pituitary. We also found that serum LH level was declined at between 12 and 60 min with the lowest level at 24 min after NMS injection. The decreased LH secretion induced by NMS could be completely abolished by pretreatment with melanocortin receptor-4 antagonist SHU9119, while a signal injection of 1.0 nM SHU9119 per se did not affect the serum LH level. Real time quantitative RT-PCR results showed that the expression of GnRH and LH mRNAs were down-regulated by NMS treatment, but their reduction was restored to normal level by SHU9119 treatments. The expression of NMU2R and PR mRNAs were up-regulated by NMS treatment, but their effects were blocked by SHU9119 treatments. The expression of the estrogen receptor mRNA in the pig hypothalamus and pituitary was unchanged under the NMS and SHU9119+NMS treatments. In summary, all results suggest that the inhibitory effect of NMS on LH is at least in part through its receptor NMU2R and mediated via MSH neurons in hypothalamus-pituitary axis of ovariectomized pigs.

Sir Bernard Katz: 26 March 1911 - 20 April 2003.

Sir Bernard Katz established the cellular basis of synaptic transmission at the neuromuscular junction, the contact point between nerve and muscle. With his death, we lost one of the most distinguished biophysicists of our time. He laid the foundations for our understanding of almost every aspect of synaptic transmission. Bernard Katz revealed the existence of key molecules and formally described their interaction. With the benefit of his almost magical intuition, he formulated hypotheses that are now recognized as facts. During his career he pioneered research in three areas. He and Alan Hodgkin elucidated the ionic basis of the action potential overshoot, as formulated in the sodium hypothesis; he unravelled the biophysical mechanisms that generate the endplate potential; and he clarified mechanisms of transmitter release, as detailed in the quantal hypothesis and the vesicle hypothesis. In particular his work on the neuromuscular junction influenced and led several generations of neurophysiologists, and it continues to do so even though research focus has shifted to synapses in the central nervous system. Bernard Katz (or BK, as he was known to colleagues and students) trained several generations of young investigators who have been inspired by his hypotheses, by his impeccable thoroughness as an investigator, and by the straightforward, unpretentious style of his presentations - though some have been dismayed by his occasional unapproachability or his unforgiving nature when confronted with others' mistakes! Perhaps his most valuable and enduring legacy to collaborators and students is that, when data are difficult to interpret and we see only a faint glimmer of light at the end of a long tunnel, we ask ourselves, 'What would BK do now?'

Neuromedin U receptor 1 expression in the rat endocrine pancreas and evidence suggesting neuromedin U suppressive effect on insulin secretion from isolated rat pancreatic islets.

Neuromedin U (NmU) is a regulatory peptide found in significant concentrations in both the brain and gut of the rat and is named according to its ability to powerfully contract the uterus. Two types of NmU receptors were recently identified and subsequent studies evidenced NmU involvement in the regulation of energy homeostasis. Such a role of neuromedin U suggests that a polypeptide may also be involved in the regulation of adipoinsular axis function. Therefore in the present study we examined the expression of NmU receptors in pancreatic islets using RT-PCR and Western blotting analysis. We also investigated the role of NmU in regulation of insulin secretion in vitro using isolated pancreatic islets. We have confirmed that NmUR1 but not NmUR2 is specifically expressed in isolated rat pancreatic islets. In all tested doses (1, 10, 100 nmol/l) NmU dose- dependently decreased insulin output by isolated pancreatic islets. These inhibitory effects of NmU on insulin secretion may suggest the involvement of NmU in regulating the pancreatic branch of adipoinsular axis function. Thus, NmU can be included in that group of anorectic peptides, which are also involved in the regulation of insulin secretion.

Importance of P-cadherin, beta-catenin, and Wnt5a/frizzled for progression of melanocytic tumors and prognosis in cutaneous melanoma.

PURPOSE: It has been proposed that melanoma cells shift from E-cadherin to N-cadherin expression during tumor development, and recent gene profiling has shown increased expression of Wnt5a/Frizzled in aggressive melanomas possibly by interactions with beta-catenin. We therefore wanted to investigate the role of cadherin subtypes, beta-catenin, and Wnt5a/Frizzled in melanocytic tumors, with focus on prognosis in nodular melanomas. EXPERIMENTAL DESIGN: The immunohistochemical expression of E-cadherin, N-cadherin, P-cadherin, beta-catenin, and Wnt5a/Frizzled was examined using tissue microarrays of 312 melanocytic tumors. RESULTS: Cytoplasmic expression of P-cadherin was associated with increasing tumor thickness (P=0.005) and level of invasion (P=0.019), whereas membranous staining was associated with thinner (P=0.012) and more superficial (P=0.018) tumors. Increased cytoplasmic P-cadherin was associated with reduced survival (P=0.047). Lack of nuclear beta-catenin expression was related to increased tumor thickness (P=0.002) and poor patient survival in univariate (P=0.0072) and multivariate (P=0.004) analyses. Membranous expression of N-cadherin was significantly increased from primary tumors to metastatic lesions, whereas E-cadherin staining tended to be decreased. Wnt5a and its receptor Frizzled were highly coexpressed, and nuclear expression of both markers was significantly reduced from benign nevi to melanomas, with a shift from nuclear to cytoplasmic expression in malignant tumors. In addition, Wnt5a expression was significantly associated with nuclear beta-catenin expression. CONCLUSIONS: Alterations in the expression and subcellular localization of cell adhesion markers are important in the development and progression of melanocytic tumors, and strong cytoplasmic P-cadherin expression and loss of nuclear beta-catenin staining were associated with aggressive melanoma behavior and reduced patient survival.

[Neurological explorations in Nuclear Medicine. A Spanish survey]. / Exploraciones neurológicas en Medicina Nuclear. Encuesta española.

A representative Nuclear Medicine Service in Spain has two or three gammacameras. It performs neurologic studies one or two days at week wich account 2,1% of total workload. Brain Perfusion SPECT, specially in cognitive disorders, is the most frequent application. Neurooncology has a lower but established relevance. Neuroreceptors imaging are increasing in the last months. Emission tomography is obtained using a double-headed camera fitted with high-resolution parallel hole collimators and a half an hour total acquisition time. Datasets are reconstructed by filtered backprojection with Butterworth or Metz filtres. Images are visually interpreted with comparison to MRI and/or CT findings.

Wnt and frizzled receptors as potential targets for immunotherapy in head and neck squamous cell carcinomas.

The diverse receptor-ligand pairs of the Wnt and frizzled (Fz) families play important roles during embryonic development, and thus may be overexpressed in cancers that arise from immature cells. Hence, we investigated the expression and function of five Wnt (Wnt-1, 5a, 7a, 10b, 13) and two Fz (Fz-2, 5) genes in 10 head and neck squamous carcinoma cell lines (HNSCC). In comparison to normal bronchial or oral epithelial cells, all the HNSCC had markedly increased mRNA levels of Wnt-1, 7a, 10b, and 13, as well as Fz-2. Moreover, the levels of Wnt-1, 10b, and Fz-2 proteins were also markedly increased in HNSCC, relative to normal epithelial cells. Treatment of one HNSCC cell line (SNU 1076) with anti-Wnt-1 antibodies reduced the activity of the Wnt/Fz dependent transcription factor LEF/TCF, and diminished the expression of cyclin D1 and beta-catenin proteins. Blocking Wnt-1 signaling also inhibited proliferation and induced apoptosis in these cells. These results show that HNSCC cell lines often overexpress one or more Wnt and Fz genes, and suggest that the growth and survival of a subset of HNSCC may depend on the Wnt/Fz pathway. Hence, the Wnt and Fz receptors may be possible targets for immunotherapy therapy of this common cancer.

Properties of excitatory amino acid transport in the human U373 astrocytoma cell line.

In this study, we describe the presence of Na(+)-dependent high-affinity L-glutamate transport activity in the human U373 astrocytoma cell line. U373 cells exhibited a robust accumulation of L-glutamate which was predominantly (85%) extracellular Na(+)-dependent. Kinetic analysis of this transport activity revealed that the uptake followed first-order Michaelis-Menten kinetics and was high-affinity in nature. The kinetic parameters estimated by Eadie-Hofstee transformation of the saturable uptake were 37.3 +/- 5.1 microM for K(m) and 0.13 +/- 0.02 nmol min-1 mg-1 protein for Vmax. A total of 14 known inhibitors of high-affinity L-glutamate transport were examined for their abilities to inhibit L-glutamate uptake by U373 cells. Three compounds, kainate (KA), dihydrokainate (DHK) and alpha-aminoadipic acid produced less than 30% inhibition at 1 mM. The lack of effect of both KA and DHK indicates that the predominant astroglial L-glutamate transporter EAAT2 (excitatory amino acid transporter 2) does not contribute to the uptake activity present in these cells. The rank order of inhibitory potency for the remaining 11 compounds tested was L-cysteine sulphinate = L-CCG-III = L-cysteate = L-aspartate = threo-beta-hydroxyaspartate > trans-PDC > D-aspartate = MPDC > beta-glutamate > L-CCG-IV = L-aspartate-beta-hydroxamate. Pre-treatment of U373 cells with phorbol ester for 30 min resulted in a 56% decrease in L-glutamate uptake and this effect was blocked in a concentration-dependent manner by the PKC inhibitor bisindolylmaleimide I. Expression of L-glutamate transporters by U373 cells was examined by reverse transcriptase polymerase chain reaction (RT-PCR) and Western analysis. Transcripts for both the EAAT1 and EAAT3 transporter subtypes were detected but not for EAATs 2, 4, and 5. Immunoblot analysis confirmed the presence of EAAT3 protein, however, we were unable to detect EAAT1 protein. In conclusion, the Na(+)-dependent high-affinity L-glutamate transport into human U373 astrocytoma cells appears to be mediated predominantly by the EAAT3 subtype.

Regulatory peptide receptors as molecular targets for cancer diagnosis and therapy.

Regulatory peptides are small, readily diffusable and potent natural substances with a wide spectrum of receptor-mediated actions in humans. High affinity receptors for regulatory peptides such as somatostatin, substance P, vasoactive intestinal peptides, and cholecystokinin can be overexpressed in several human diseases, in particular in neoplasms, and represent therefore new molecular targets for cancer diagnosis and therapy. The availability of suitable regulatory peptide radioligands, which can be labeled with radioactive iodine or indium, makes peptide receptor scintigraphy a particularly useful new in vivo diagnostic tool, as seen with the example of somatostatin receptor scintigraphy (Octreoscan). The present article reviews the current in vitro data on regulatory peptide receptor expression in normal and diseased tissues, which represent the basis for the in vivo application of these molecules in nuclear medicine.

Subpopulations of neonatal rat sensory neurons express functional neurotransmitter receptors which elevate intracellular calcium.

We have attempted to identify which subpopulations of rat sensory neurons possess functional neurotransmitter receptors which elevate the free concentration of intracellular calcium. Subpopulations of sensory neurons were identified using three accepted criteria: (i) the distribution and proportion of neurons with differing somatic diameters; (ii) the expression of substance P-like immunoreactivity; and (iii) the responsiveness of each neuron to capsaicin. The total neuronal population was primarily grouped into three classes according to somatic diameter and defined as small- (< 17 microns), intermediate- (17-25 microns) and large- (> 25 microns) sized neurons. It was not possible to distinguish between small and intermediate-sized neurons since a similar percentage of each class expressed substance P-like immunoreactivity or sensitivity to capsaicin. Large-sized neurons did not possess these characteristics and, therefore, represented a distinct neuronal population. In single, intact neurons of differing diameter, the ability of a variety of receptor agonists to elevate the free concentration of intracellular calcium was determined using the calcium-sensitive indicator, Fura-2. Local application of capsaicin, adenosine, bradykinin, ATP and substance P elevated the resting level of the free concentration of intracellular calcium in small and intermediate-sized neurons. The large-sized neurons were unresponsive to these receptor agonists with the exception of ATP. The response to ATP was relatively transient in nature and did not differ between neurons of differing somatic diameter.(ABSTRACT TRUNCATED AT 250 WORDS)

Neural grafting to ischemic CA1 lesions in the rat hippocampus: an autoradiographic study.

Fetal hippocampal neurons were stereotaxically transplanted to five-day-old ischemic CA1 lesions in adult rat hippocampi. The recipient brains were examined 14 or 100 days later. The grafts survived well, and transplanted cells usually formed clusters in the host CA1 subfield. In vitro receptor autoradiography was employed to map the following receptors, the ligands indicated in parentheses being used for labeling: muscarinic cholinergic ([3H]quinuclidinyl benzilate), adenosine A1 ([3H]cyclohexyladenosine), kainate ([3H]kainic acid), spirodecanone ([3H]spiperone), opioid ([3H]naloxone), and GABAA ([3H]muscimol). The receptor autoradiographic technique showed significant binding of the six ligands in all hippocampal grafts two weeks after transplantation. One hundred days following transplantation, almost all receptors, especially muscarinic cholinergic, adenosine A1 and opioid receptor bindings in grafts, had significantly increased compared to bindings two weeks after transplantation. At this time, kainate and muscarinic cholinergic receptors in grafts had increased up to the near normal level of the CA1 in the hippocampus. Interestingly, adenosine A1 receptors in the grafted side had significantly increased not only in the CA1 but also in the stratum oriens of the CA3 compared with that in the non-grafted side. The increase of [3H]quinuclidinyl benzilate binding corresponded well with the innervation of acetylcholinesterase-positive fibers at 100 days after grafting. These results demonstrate that the transplanted neurons, which showed both pre- and post-synaptic autoradiographic markers in the ischemic CA1 lesions, are able to develop their properties and express the nature of normal hippocampal neurons.

Evidence for further heterogeneity of the receptors for neuropeptide-Y and peptide-YY in tumor cell lines derived from neural crest.

The expression and structure of the receptors for neuropeptide-Y (NPY) and peptide-YY (PYY) were studied in 16 human and rodent tumor cell lines derived from the neural crest by ligand binding and cross-linking techniques using [125I]Bolton-Hunter-NPY, [125I]PYY, and various forms of monoiodinated NPY and PYY. Although NPY-binding sites were observed in most of the tumor cells, PYY-binding sites were found only on the human neuroblastoma cell lines SMS-MSN, SMS-KAN, SK-N-MC, and MC-IXC and the human Ewing's sarcoma cell line SK-ES. The differential labeling of the NPY/PYY receptors on these cell lines suggests that the NPY/PYY receptors are more heterogeneous than previously described as the Y1, Y2, and Y3 receptor subtypes. Cross-linking studies demonstrate that the Y1 and Y2 receptors for NPY/PYY are structurally different (mol wt, 70 and 50 kilodaltons, respectively) and that the 70- and 50-kilodalton receptor proteins are coexpressed in certain tumor cell lines. This could explain at least in part why cell lines show a relative specificity for Y1/Y2 classification, observed as the inhibition by both C-terminal fragments and Y1-specific analogs on the NPY/PYY binding to membrane receptors. Collectively, the present study suggests further heterogeneity of the NPY/PYY receptors and the existence of multiple receptor proteins in the tumor cell lines derived from the neural crest.

Somatostatin receptor imaging in the diagnosis and treatment of neuroendocrine tumors.

Somatostatin analogs are used in the control of hormonal hypersecretion and tumor growth of patients with acromegaly, islet cell carcinomas and carcinoids. Recently we showed that somatostatin receptor positive tumors can be visualized in vivo after the administration of radioactive isotope-labelled somatostatin analogs. Receptor imaging was positive in 18/21 islet cell tumors, 30/31 carcinoids, 26/28 paragangliomas, 9/14 medullary thyroid carcinomas, 5/7 small cell lung cancers, 6/7 neuroblastomas, 38/49 primary breast cancers, and 0/18 pancreatic adenocarcinomas. Also 11/11 meningiomas, 4/4 astrocytomas and 0/3 glioblastomas could be visualized. Somatostatin receptor imaging is an easy, harmless and painless diagnostic method. It is an in vivo method for the recognition of neuroendocrine cancers. It localizes multiple and/or metastatic tumors, predicts the successful control of hormonal hypersecretion by octreotide and seems of prognostic value in certain types of cancer. This scintigraphic method might help in patient selection for clinical trials with somatostatin analogs in the treatment of neuroendocrine cancers.

Immunohistochemical localization of receptors for vasoactive intestinal peptide and substance P in human trachea.

BACKGROUND: Tissue sections of human cervical trachea were processed for immunohistochemical demonstration of receptors for substance P [using an anti-SP anti-idiotypic antiserum directed toward the ligand binding site of the receptor (Couraud J-Y, Escher ED, Regoli D, Imhof V, Rossignol B, Pradelles P. Anti-substance P anti-idiotypic antibodies: Characterization and biological activities. J Biol Chem 1985;260:9461-9; Couraud J-Y, Maillet S, Grassi J, Frobert Y, Pradelles P. Characterization and properties of anti-substance P antiidiotypic antibodies. Methods Enzymol 1989; 178:275-300)] and vasoactive intestinal peptide (VIP; utilizing a monoclonal antibody toward VIP receptors of an adenocarcinoma cell line (Pichon J, Hirn M, Muller J-M, Mangeat P, Marvaldi J. Anticell surface monoclonal antibodies which antagonize the action of VIP in a human adenocarcinoma cell line (HT29). EMBO J 1983;2:1017-22)], respectively. Mucus cells of the submucosal glands (identified by periodic acid Schiff staining) and neuroendocrine cells of the respiratory epithelium (identified by immunoreactivity to protein gene product 9.5) displayed intense VIP receptor-immunoreactivity. Other tissue components known to respond to exogenously administered VIP, e.g., trachealis muscle, lacked VIP receptor-immunoreactivity, indicating that the monoclonal antibody did not label all receptor subtypes. In accordance with the known pharmacological actions of substance P upon the airways, the anti-substance P receptor antibody labeled the trachealis muscle, submucosal glands, and respiratory epithelium, predominantly at the luminal aspect. Since substance P as well as the structurally related tachykinin, neurokinin A, competed with the anti-receptor antibody in binding to the tissue section, it is likely that both NK-1 and NK-2 receptor subtypes were labeled. The present histochemical approach to localize peptide receptors in the trachea allowed precise analysis of distribution unreached by previous studies using autoradiography. Together with pharmacological data, these morphological findings contribute to the understanding of the sequences of events evoked by the neuropeptides, substance P and VIP, in the human trachea.

In vitro detection of somatostatin receptors in human tumors.

Somatostatin receptors (SSR) have been identified in membrane homogenates or tissue sections from several hundred human tumors. SSR have been found in most neuroendocrine tumors, ie, growth hormone (GH)- and thyrotropin (TSH)-producing pituitary tumors, endocrine gastroenteropancreatic (GEP) tumors, paragangliomas, pheochromocytomas, medullary thyroid carcinomas (MTC), and small-cell lung carcinomas. SSR have also been found in the majority of malignant lymphomas, in several brain tumors (all meningiomas, most astrocytomas), and in breast tumors. The majority of tumors expressing SSR are rather differentiated, eg, astrocytomas in contrast to glioblastomas, but exceptions such as high-grade malignant lymphomas do exist. An inverse relationship exists between SSR and receptors for epidermal growth factor in lung tumors, glial tumors, and most breast tumors, whereas meningiomas express both receptors simultaneously. A minority of tumors such as ovarian tumors, MTC, and insulinomas express a subtype of SSR characterized by low affinity for the octapeptide SS analogue, octreotide. The function of SSR in human tumors differs according to tumor type; SSR in pituitary and GEP tumors mediate hormone secretion inhibition and possibly have some antiproliferative effects. However, in meningiomas, activation of SSR inhibits forskolin-stimulated adenylate cyclase activity and weakly stimulates proliferation. Although SSR seem to mediate antiproliferative effects in animal models and cell lines of lymphomas and breast and lung tumors, such an effect has not yet been convincingly documented in human primary tumors.(ABSTRACT TRUNCATED AT 250 WORDS)

Brain corticotropin-releasing hormone receptors on neurons and astrocytes.

UNLABELLED: Corticotropin-releasing hormone (CRH) exerts many potent effects within brain and is considered an important brain neuroregulator. CRH acts via receptors that are widely distributed throughout brain which exhibits highest CRH receptor concentrations in extrahypothalamic regions. We have previously characterized CRH receptors in heterogeneous extrahypothalamic forebrain cell cultures consisting of neurons and glia, and have shown them to exhibit similar kinetic and pharmacological characteristics as CRH receptors in pituitary and in situ brain. However, it is not known whether CRH receptors are present on neurons, glia or both. We tested the hypothesis that CRH receptors are present on neurons in extrahypothalamic forebrain cell cultures derived from day 17-18 fetal rats by characterizing receptors in predominantly neuronal (N), glial/astrocytic (G) cultures and mixed (M) cultures. Mean CRH receptor concentrations (fmol/mg protein) in N (10.4), G (9.4), and M (9.8) cultures were similar. Following Scatchard analyses derived from competition curves, all cell populations exhibited similar mean high-affinity/low-capacity (Kd = 1.0-1.9 nM; Bmax = 183-388 fmol/mg protein) and low-affinity/high-capacity (Kd = 92-104 nM; Bmax = 2034-5008 fmol/mg protein) classes of binding sites. IN CONCLUSION: (1) Neurons and astrocytes in fetal extrahypothalamic brain cell cultures contain CRH receptors which exhibit similar concentrations and similar kinetic characteristics. (2) These observations suggest that biological effects of CRH in brain could be mediated via actions on neurons and/or glial astrocytes.

[125I]iodomelatonin binding sites in the chicken brain: diurnal variation and effect of melatonin injection or pinealectomy.

The diurnal variation of [125I]iodomelatonin binding sites in the brain and serum melatonin levels were studied at 4-hour intervals under a 12 h:12 h light:dark cycle in 5-week-old chicks. There was a diurnal rhythm of [125I]iodomelatonin binding capacities in brain membrane preparations with the peak in the light period and the trough in the dark period. However, a diurnal variation with high levels in the dark and low levels in the light was observed in serum melatonin concentrations. The binding capacity of [125I]iodomelatonin was inversely related to the serum melatonin level. Four-week-old chicks were injected with melatonin (1 mg melatonin/kg; i.p.) twice a day (shortly after the light onset and shortly before light offset) for 3 weeks. When chickens were killed at 7 weeks, there was no diurnal rhythm of melatonin in the circulation. In addition, melatonin injection also abolished the diurnal variation of [125I]iodomelatonin binding sites in brain membrane preparations in these animals. This was achieved by decreasing the binding capacity in the light period. Pinealectomy in 2-day-old chickens increased the density of [125I]iodomelatonin binding sites in the brain membrane preparations both at midlight and middark when the chicks were later sacrificed at 6 weeks of age, although the rhythm of binding sites persisted. It is suggested that the [125I]iodomelatonin binding capacity in the chicken brain is regulated by endogenous melatonin and influenced by exogenous melatonin. In addition, the rhythmicity of [125I]iodomelatonin binding sites in the chicken brain is affected by, but not dependent on, pineal melatonin.

Gastrin-releasing peptide: in vivo and in vitro growth effects on an acinar pancreatic carcinoma.

The mammalian gastrin-releasing-peptide (GRP) and its structural amphibian analogue, bombesin, are known to be trophic factors for the normal exocrine pancreas. This work investigates the possible role of GRP in the growth of an acinar pancreatic cancer transplanted to the rat and in primary tumor cell cultures. Moreover, this adenocarcinoma was tested for its content of specific bombesin/GRP receptors by using autoradiographic technics and in vitro binding assays with tumor cells. In Lewis rats bearing the pancreatic carcinoma transplanted s.c. in the scapular region, chronic administration of GRP at the dose 30 micrograms/kg/day for 15 successive days significantly increased the tumor volume, the final tumor weight, and amylase, protein, RNA and DNA contents. Autoradiographic studies showed that tumor tissue was GRP receptor positive with a high density. The biochemical characterization indicated that receptor positive tumor tissue had saturable and high affinity receptors with pharmacological specificity for GRP and its bioactive analogues. In primary tumor cell cultures, GRP increased the incorporation of [3H] thymidine in DNA in a dose- and time-dependent manner. There was a good correlation between the ability of GRP and its COOH terminal analogues to elicit DNA synthesis and their affinity for 125I-GRP binding sites. These results from in vivo and in vitro experiments demonstrated that GRP induces growth of pancreatic carcinoma by acting directly on specific membrane receptors present on the tumor cells.