bluuebunny/crossref_metadata_2025 huggingface dataset:
Neuron‐specific growth cone properties as seen in identified neurons of Helisoma
Authors: Haydon P.G. , Cohan C.S. , McCobb D.P. , Miller H.R. , Kater S.B.
Published: January 2005

By using cell‐cultured identified neurons of the snail Helisoma, we demonstrate that the growth cones of different neurons are intrinsically different from one another in terms of their structure, behavior, and response to environmental signals. Structurally neuron 5 has a greater number of filopodia per growth cone, has shorter filopodia, and has a smaller interfilopodial distance than neuron 19. Behaviorally, the growth cones of neuron 5 advance over the substratum at a faster rate than those of neuron 19; and the growth cones of neuron 19, but not of neuron 5, respond to the presence of serotonin in their environment by retracting their filopodia.

In addition to such intrinsic differences between the growth cones of different neurons, we also demonstrate that the separate growth cones of a single neuron, while having identical properties, can act independently of one another. Focal application of serotonin to a growth cone causes only a localized retraction of that growth cones' filopodia. Other growth cones that are attached to the same neuron but that are not exposed to serotonin retain their normal structural features.
Guidance of neurite outgrowth by pathways of substratum‐adsorbed laminin
Authors: Hammarback J.A. , Palm S.L. , Furcht L.T. , Letourneau P.C.
Published: January 2005

Neurite outgrowth is guided by narrow pathways of bioactive laminin. These pathways are created by ultraviolet light irradiation of laminin‐coated coverslips masked with electron microscope grids. Patterned outgrowth of neurites is independent of gross mechanical guidance and guidance caused by substrate limitation. Cells on unirradiated laminin are less readily displaced by shear forces than cells on irradiated laminin. This study suggests that ultraviolet light alters the adhesive properties of laminin and that differential cell‐substratum adhesion may guide extending neurites on the purified naturally occurring substance, laminin.
Taurine deficiency in the developing cat: Persistence of the cerebellar external granule cell layer
Authors: Sturman J.A. , Moretz R.C. , French J.H. , Wisniewski H.M.
Published: January 2005

Dietary taurine deprivation adversely affects feline pregnancy and is associated with the frequent occurrence of fetal resorption, abortion, stillbirth, and low birthweight of live kittens at term. Taurine‐deprived, live‐born kittens have a poor postnatal survival rate and grow less well than kittens from taurine‐supplemented queens. The postnatal dietary taurine intake of such kittens is reduced if they are nursed by their biologic mothers; the concentration of taurine in milk of taurine‐deprived mothers is less than 10% of that in milk from taurine‐supplemented queens. Surviving kittens from taurine‐deprived mothers exhibit a constellation of neurological abnormalities (abnormal hind leg development, a peculiar gait characterized by excessive abduction and paresis, and thoracic kyphosis readily visible by X‐ray). These findings suggest the presence of a developmental cerebellar deficit. Histological examination of the pre‐ and postnatally taurine‐deprived kitten's cerebellum reveals a persistence of the external granule cell layer, which was confirmed by electron‐microscopic examination. Numerous mitotic figures are present in the cells in the external granule cell layer of the cerebellum of kittens born from and nursed by taurine‐deprived queens, but not in those from taurine‐supplemented mothers. These findings suggest a maturational delay.
Immunohistochemical localization of myelin basic protein in the developing optic system of trout
Authors: Jeserich G. , Jacque C.
Published: January 2005

Immunofluorescent localization of basic protein (BP) in the CNS of trout was performed using a heterologous antiserum raised against human BP. Bright specific fluorescence was confined to the myelin sheath of axons in most regions of the brain, whereas neuronal pericarya and dendrites as well as glial cells were entirely negative. In the tectum immunofluorescence was characteristically distributed in two rows of horizontally aligned patches, most evidently reflecting the orderly array of myelinated fiber bundles in the stratum opticum and stratum album. During development of the retino‐tectal pathway, the appearance of BP followed a distinct rostro‐caudal gradient firstly reaching the anterior ventral border of the tectum by larval stage 34. Furthermore, on cross sections through the optic nerve a specifically organized pattern of myelin formation was revealed by immunohistochemistry, which strikingly coincided with the pattern of fiber outgrowth.
Quantitation of the in vitro neuroblastoma response to exogenous, purified gangliosides
Authors: Leskawa K.C. , Hogan E.L.
Published: January 2005

Individual ganglioside species (possessing the gangliotetrose oligosaccharide) were purified from bovine brain gray matter and applied in varying concentrations to the culture medium of mouse neuroblastoma cells (N2A) in vitro. After 48 hr of incubation, the cells were stained, and the neuritogenic response quantitated with a video analysis system, employing a program to measure three parameters of neuroblastoma differentiation: neurites per cell (sprouting), neurite length (extension), and degree of neurite branching (arborization). All the individual gangliosides tested promoted neurite extension in a dose‐dependent fashion. Asialogangliosides (“neutral” glycosphingolipids) were without effect, which suggests that sialic acid (N‐acetylneuraminic acid) is necessary to elicit this cellular response. With increasing concentrations of GM1 (5 to 500 μg/ml), the average cellular neurite length increased significantly, whereas the number of neurites per cell decreased. With the trisialoganglioside GT1b, neurite length did not increase to the extent seen with GM1, but an increase in the number of neurites per cell (sprouting) and branch points per neurite (arborization) was observed. These results suggest that the in vitro neuronal response to exogenous gangliosides may combine specific responses to individual species making up the total.
Asymmetric molecular forms of acetylcholinesterase in mammalian skeletal muscles
Authors: Sketelj J. , Brzin M.
Published: January 2005

Velocity sedimentation analysis of acetylcholinesterase (AChE) molecular forms was performed separately in endplate‐rich and endplate‐free regions of the diaphargm muscle of the rat, guinea pig, rabbit, dog, and pig, and in mm. erectores trunci and m. vastus lateralis in man. Several high‐ionic‐strength media were first tested to achieve better solubilization of AChE from rat muscles than by the usual 1 M NaCl‐ Triton X‐100 medium. Ninety‐five percent of the rat diaphragm was solubilized in a single extraction step by medium containing 1 M lithium chloride instead of NaCl. Homologous molecular forms of AChE were found in all species. The asymmetric forms were invariably present in the endplate regions of muscles but their activity in endplate‐free regions was much lower than in endplate regions in all investigated mammals except in man. Essentially the same pattern of AChE molecular forms was present in both regions in human muscles. High extrajunctional activity of the asymmetric forms makes human muscles similar to immature rodent muscles in vivo and in culture. The pattern of AChE molecular forms in the endplate region of the diaphragm in senile 24‐month‐old rats was not significantly different from that in 3‐month‐old animals. The persistence of the asymmetric AChE forms in the diaphragm of senile rats suggests that neuromuscular interactions do not become deficient with age in this muscle.
Excitatory effect of amino acids on identified neuron R14 of Aplysia. I. Glycine‐induced depolarization and its ionic mechanism
Authors: Ichinose M. , Sawada M. , Maeno T.
Published: January 2005

The ionic mechanism of the membrane effect of glycine on identified neuron R14 of Aplysia was investigated with conventional intracellular recording and voltage‐clamp techniques. Both localized and bath applications of glycine markedly depolarize R14. Bath‐applied glycine induced an inward current that gradually reached a maximum and remained at that level until glycine was washed out. Displacement of the holding potential from –46 to –121 mV increased the inward current. The extrapolated reversal potential was +38.6 mV. Reduction of [Na+]o reversibly decreased the inward current. Alterations of [K+]o, [Cl−]o, and [Ca2+]o, as well as bath‐applied ouabain and sodium cyanide, did not affect the inward current. These results suggest that glycine can induce an Na+ current and that the glycine‐induced inward current does not reflect an active uptake by an Na+‐coupled transport system of glycine into the neuron.
Development of mouse spinal cord in tissue culture: IV. Effects of embryonic extracts on neuron formation and migration
Authors: Houle J. D. , Fedoroff Sergery
Published: January 2005

Possible influences upon patterns of neurogenesis expressed in vitro were examined quantitatively by the use of microfragment cultures of embryonic day 10 mouse neural tube. Crude extracts were prepared either from whole embryos (day 13 or 15 of gestation) or from embryonic brains (day 18 of gestation) and added to the culture medium for the first 10 days of culturing. Neuronal outgrowth zones surrounding individual microfragments were reduced in area (indicating restricted neuronal migration) and in number of neurons present (indicating restricted production of neurons) following treatment with either of the extracts. The severity of reductions observed were related to the developmental age of embryonic tissue used for preparing the extract, as greatest reduction resulted from addition of embryonic day 18 brain extracts and to concentration employed, higher doses further restricting neuronal outgrowth. By increasing the concentrations of extract the proportional number of large‐sized neurons forming the outgrowth zones became greater relative to the small neuron contribution, indicating an enhanced survival for this neuronal population. The formation and migration of astroglial precursor cells was not affected by the addition of any of the extracts.

The number of neurons remaining within the original portions of neural tube microfragments was not significantly altered following culturing in the presence of embryonic extract. This suggested that the reduction in neuron number in the outgrowth zone actually reflected a decreased neuron production and was not simply the result of a retention of neurons within the remaining portion of the microfragment
.

The results suggest the presence of substances within mouse embryos that have regulatory effects on aspects of development of the central nervous system. Indications are that survival and maturation of postmitotic neuroblasts are promoted in vitro while the formation of additional neuronal progenitor cells may be partially inhibited by the addition of embryonic mouse extracts to the medium. We propose that an endogenous negative feedback mechanism may be invloved in the coordination of patterns of neurogenesis.
Identification of a neuroregulated phosphoprotein in skeletal muscle as the regulatory subunit of cyclic AMP‐dependent protein kinase II
Authors: Squinto S. P. , McLane J. A. , Held I. R.
Published: January 2005

When soluble proteins in cytosolic fractions of rat soleus muscles are 32P‐phosphorylated in vitro by an ATP:protein phosphotransferase reaction, the major substrate is a 56‐kilodalton (56K) protein. As we have also reported previously, the onset and development of increased 32P‐phosphorylation of this 56K protein, which are observed after the soleus is denervated, temporally correlate with the denervation period and length of the distal nerve stump [Held et al, 1983]. Conclusive evidence which identifies this neuroregulated muscle protein as the regulatory subunit of cyclic AMP‐dependent protein kinase type II (R‐II) is presented in this paper. The 56K soleus protein and purified bovine heart R‐II were 32P‐phosphorylated and subjected to limited proteolysis with bovine pancreas trypsin. After resolution of the generated 32P‐phosphopeptides by SDS slab PAGE and visualization by autoradiography, no tryptic products were observed from the 56K soleus protein which were not also produced by proteolysis of the purified R‐II. These tryptic phosphopeptides included 39, 16.5, and 12K fragments which retained the autophosphorylation site of R‐II. After denervation, the 32P‐phosphorylation of the 56K soleus protein and of the 39K tryptic peptide product were comparably increased. The identification of the neuroregulated 56K soleus protein as R‐II was also confirmed by Western blotting with a specific anti‐R‐II sera. Taken together, our results demonstrate that the previously observed neuroregulation of the 32P‐phosphorylation of the 56K soleus protein is identifiable with some alteration which affects the intramolecular 32P‐autophosphorylation of R‐II.
Amino acid incorporation during morphine intoxication. II: Electrophoretic separation of extracellular proteins from cerebral hemisphere slices and astroglia‐enriched primary cultures
Authors: Hansson E. , Rönnbäck L.
Published: January 2005

Radioactively labeled proteins were identified in a broad molecular weight range in the incubation medium of rat cerebral hemisphere slice preparations or primary cultures after incubation with radioactive valine. Morphine chloride caused an increase in labeled media proteins with MW ˜40,000 and 65,000 in both preparations, whereas a fraction with MW, ˜80,000 decreased in amount. In the culture preparation a MW ˜15,000 fraction also decreased after morphine treatment. The effects on the MW 40,000 and 65,000 fractions by morphine could not be blocked by naloxone hydrochloride, an opiate antagonist. Labeled media proteins from primary cultures obtained from various brain regions showed many similarties. After 10−5 or 10−6 M morphine, protein fractions with MW ˜15,000, 25,000, 40,000, 65,000–70,000, 80,000, and 100,000 changed with increases or decreases in the various morphine concentrations. The 3H‐labeling of the MW ˜40,000 fraction increased in all cultures, where it decreased.

In conclusion, proteins synthesized within cells in brain slices or in astrogliaenriched cultures are released or secreted into the incubation media. Some protein fractions are affected in different directions by morphine. The effects were not correlated with the presence of classical opiate receptors on the cells. This was so even though more changes in media proteins occurred in astroglial cultures from opiate receptor‐rich brain regions than in similar cultures from opiate receptor‐poor regions. The possible significance of proteins synthesized in astroglial cells and released extracellulary in opiate action is discussed.
Effect of morphine‐induced cortical excitation on spinal sensory transmission
Authors: Hernandez A. , Soto‐Moyano R.
Published: January 2005

Bioelectrical responses evoked in the ventrolateral funiculus (VLF) of the spinal cord by electrical stimulation of the contralateral hind limb were studied following topical application of 1% morphine solution to the somatosensory SI area of the rat cerebral cortex. After morphine, a typical pattern was observed in the electrocorticogram, characterized by the appearance of rhythmic spiking activity. Time‐related with each cortical spike, a significant reduction in the amplitude of VLF responses was observed. It is concluded that cortical excitation induced by morphine generates descending influences having the ability to inhibit spinal sensory transmission.
Abnormal visual cortex development in the kitten associated with maternal dietary taurine deprivation
Authors: Palackal T. , Moretz R. , Wisniewski H. , Sturman J. A.
Published: January 2005

We have examined the visual cortex of newborn and 8‐week‐old kittens born to mothers consuming a taurine‐supplemented or taurine‐free diet using the rapid Golgi technique. Kittens from taurine‐supplemented mothers exhibited normal development of the visual cortex. Kittens from taurine‐depleted mothers showed striking differences. In newborn kittens, neuroblasts are aggregated both at the ventricular and pial zones, having failed to migrate and differentiate normally. Eight weeks after birth, only few pyramidal and nonpyramidal neurons are found. Those present have heavily spined dendritic processes indicative of poor arborization. Protoplasmic astrocytes are represented by undifferentiated cellular masses. The taurine concentration in the cortex of such kittens is four‐ to fivefold smaller than in kittens from taurine‐supplemented mothers.

These results suggest that normal concentrations of taurine in visual cortex, as well as cerebellum, are required for normal ontogeny of neurons. Once such deficits are established in the prenatal and immediate postnatal period, they result in permanent abnormalities. These findings have clear implications for vegetarian women who intend to have children, since virtually no taurine is present in plants and vegetables.
Cellular effects of a brain extract factor or factors inhibiting neuroblastoma cell growth in vitro
Authors: Schengrund C.‐L. , Repman M. A.
Published: January 2005

The mechanism of inhibition of neuroblastoma cell proliferation in vitro by extract prepared from the brains of newborn mice was partially elucidated. Flow cytometric analysis of propidium iodide‐stained cells showed that a significant increase occurred in the pre‐G0/G1 cell population after exposure to the extract for 24 hr. Fluorescence microscopy indicated that the pre‐G0/G1 cells had small “micronuclei” in association with their nucleus, whereas post‐G2/M cells had enlarged propidium iodide‐staining nuclei. 3H‐thymidine incorporation was unchanged from that of control cells, in cells exposed to the extract for either 6 or 24 hr. No effect was seen on choline acetyl transferase activity or on morphology. These results suggest that the inhibitor blocks neuroblastoma cell proliferation either by inducing an uneven distribution of DNA at the time of cell division or by altering its packaging within the nucleus.
Rates of amino acid incorporation into particulate proteins in vivo and in slices of young and adult brain
Authors: Shahbazian F. M. , Jacobs M. , Lajtha A.
Published: January 2005

Incorporation of a flooding dose of valine into brain proteins was measured in vivo and in brain slices, according to subcellular fraction and to solubility. In particulate fractions in vivo in the immature brain, soluble cytosol proteins had the highest synthesis rates (2.0% per hr), followed by microsomes (1.7%), mitochondria, nuclei (1.4%), and synaptosomes (0.85%). In adult, the rates were about half of those of immature animals except for microsomes, which were the same. In young, rates of incorporation into particles in brain slices were about 80% of in vivo rates, and adult rates were 10–20% of the in vivo rates.

When brain proteins were fractionated into four groups‐isotonic soluble, hypotonic soluble, Triton X‐100 soluble, and the remainder solubilized in 0.2% SDS–in both young and adult brain in vivo, the isotonic fraction had the highest rate of protein synthesis, followed by the Triton‐soluble fraction, the SDS‐soluble fraction, and the hypotonic fraction. Also, with this method of separation in vitro rates for immature brain were 10–20% lower than in vivo rates, while in vitro adult rates were 80–90% lower.

The results indicate that the most proteins are synthesized at a higher rate in the immature brain, and that the difference between in vivo and brain slice protein synthesis is not as great in the immature as in the mature brain in all the fractions examined.
Molecular adaptations in catecholamine biosynthesis induced by cold stress and sympathectomy
Authors: Stachowiak M.K. , Fluharty S.J. , Stricker E.M. , Zigmond M.J. , Kaplan B.B.
Published: January 2005

Chronic cold stress and chemical sympathectomy are known to increase the synthesis and release of catecholamines in the adrenal medulla. Chromaffin cells adapt to altered functional requirements by increasing the synthesis of tyrosine hydroxylase (TH), the rate‐limiting enzyme in catecholamine biosynthesis. In this study, we investigated the molecular genetic mechanisms underlying these changes in enzyme activity. Estimates of TH mRNA levels were obtained by RNA dotblot analysis with a cloned TH cDNA hybridization probe. Exposure to cold produced a 4.3‐fold increase in the relative abundance of adrenomedullary TH mRNA. Increases in TH mRNA levels (90%) also were observed in the brainstem of cold‐stressed animals. The relative amount of TH synthesized in vitro in a rabbit reticulocyte cell‐free system, programmed with adrenal poly (A) +RNA, increased 4.3 times in cold‐stressed rats. Alteration in TH mRNA abundance appears to be specific, as we observed no significant difference in the levels of total RNA or poly(A) +RNA in this tissue. In addition, the relative abundance of adrenomedullary TH mRNA increased by 60% 4 days after systemic administration of the neurotoxin 6‐hydroxydopamine. This increase was transient and disappeared 2 weeks after the lesion. Changes in TH mRNA levels after cold stress or sympathectomy were eliminated by denervation of the adrenal gland. These results indicate that alterations in the relative abundance of TH mRNA mediate changes in TH activity induced by chronic stress or sympathectomy, and that these changes require an intact sympathetic input.
Use of synthetic oligodeoxynucleotide probes for the isolation of a human cholinesterase cDNA clone
Authors: Prody C. , Zevin‐Sonkin D. , Gnatt A. , Koch R. , Zisling R. , Goldberg O. , Soreq H.
Published: January 2005

Cholinesterases are serine esterases that rapidly hydrolyze the neurotransmitter acetylcholine. In humans, cholinesterases exhibit extensive polymorphism in terms of their substrate specificity, sensitivity to selective inhibitors, hydrophobicity, and cellular as well as subcellular localization. It is not yet known whether the various cholinesterase forms originate from different genes or are products of posttranscriptional and posttranslational processing. The extent to which these enzyme forms are homologous in their amino acid sequence is also not known. However, a consensus organophosphate‐binding hexapeptide sequence Phe‐Gly‐Glu‐Ser‐Ala‐Gly was found both in “true” acetylcholinesterase from the electric organ of Torpedo [McPhee‐Quigley et al: J Biol Chem 260:12185–12189, 1985] and in “pseudocholinesterase” (butyrylcholinesterase) from human serum [Lockridge: “Cholinesterases—Fundamental and Applied Aspects.” New York: de Gruyter, pp 5‐12, 1984], suggesting that this region in the protein is conserved in all cholinesterases. Based on this common sequence, we prepared synthetic oligodeoxynucleotides and used them as labeled probes to screen a cDNA library from fetal human brain mRNA, cloned in lambda gt10 phages. A cDNA clone of 770 nucleotides in length was isolated. It contains an open reading frame terminating with the sequence Ser‐Val‐Thr‐Leu‐Phe‐Gly‐Glu‐Ser‐Ala‐Gly‐Ala‐Ala, which includes the consensus hexapeptide used for designing the DNA probe. Furthermore, the sequence of this 12‐amino acid peptide is identical to the sequence reported for the organophosphate binding site of human serum pseudocholinesterase [Lockridge: “Cholinesterases—Fundamental and Applied Aspects.” New York: de Gruyter, pp 5–12, 1984]. These findings confirm that the isolated clone is indeed part of a human cholinesterase cDNA.
Neuron‐specific enolase: Complete structure of rat mRNA, multiple transcriptional start sites, and evidence suggesting post‐transcriptional control
Authors: Forss‐Petter S. , Danielson P. , Sutcliffe J.G.
Published: January 2005

The protein encoded by a randomly selected rat brain cDNA clone was identified as neuron‐specific enolase (NSE; 4.2.1.11; gamma subunit), based on homology to yeast enolase sequences and the presence of the corresponding 2.5–kb mRNA in rat brain but not in liver, kidney, or muscle tissue. The 2,222‐nucleotide NSE and mRNA sequence presented identifies a 68‐nucleotide 5′ noncoding region, a 1,302‐nucleotide open reading frame (corresponding to a primary translation product of 434 amino acids), and 852 noncoding ' bases. Evolutionary implications based on sequence comparisons to yeast enolase and non‐neuronal enolase are discussed. Primer extension analysis indicated the presence of several alternative initiation sites for transcription within 60 nucleotides on the NSE gene. The developmental onset of NSE mRNA expression correlates with the appearance of NSE protein; however, the mRNA reaches adult levels by postnatal week 3, whereas the protein continues to accumulate over the next few months, suggesting regulatory mechanisms in addition to transcriptional control.
Brain enzyme adaptation to mild normobaric intermittent hypoxia
Authors: Marzatico F. , Curti D. , Dagani F. , Tagliti M. , Benzi G.
Published: January 2005

The adaptation of repeated periods of intermittent normobaric hypoxia (oxygen:nitrogen = 10:90, 12 hr daily for 5 days) of some specific enzymatic activities related to energy metabolism has been observed in different rat brain areas (cerebral cortex, hippocampus, corpus striatum, hypothalamus, cerebellum, and medulla oblongata).

The evaluation of the maximum rate (Vmax) of the enzymes was carried out on: (1) the homogenate “in toto,” (2) the nonsynaptic mitochondrial fraction, and (3) the crude synaptosomal fraction. The adaptation to inermittent normobaric hypoxic exposure was characterized by significant modifications of some enzyme activities in the homogenate “in toto” (decrease of hexokinase activity in cerebellum), in the nonsynaptic mitochondrial fraction (increase of succinate dehydrogenase activity in corpus striatum and decrease of cytochrome oxidase activity in cererbral cortex), and, particularly, in the synaptosomal fraction (decrease of cytochrome oxidase activity in cerebral cortex, hippocampus, corpus striatum, and cerebellum, and decrease of malate dehydrogenase and lactate dehydrogenase activity in cererbellum).

The adaptation to normobaric intermittent hypoxia differs according to the brain area, subcellular fraction, and enzyme activity tested.
Chronic neurologic dysfunction and demyelination induced in lewis rats by repeated injections of encephalitogenic T‐lymphocyte lines
Authors: Vandenbark A.A. , Nilaver G. , Konat G. , Teal P. , Offner H.
Published: January 2005

Experimental autoimmune encephalomyelitis (EAE) in the Lewis rat is characteristically a monophasic paralytic disorder. Recovered rats are thereafter immune to EAE induced by injection of guinea pig basic protein (GP‐BP) in complete Freund's adjuvant (CFA), but they are still susceptible to EAE induced by an encephalitogenic T‐lymphocyte line (BP‐1). Induction of active EAE or injection of a sublethal dose of activated BP‐1 cells resulted in a monophasic episode of EAE, followed by recovery of normal neurologic function. Repeated challenges with activated BP‐1 cells, however, induced unremitting neurologic signs marked by loss of tail tonicity and incontinence, which persisted for more than 6 months. Histologically, the spinal cord of affected rats revealed attenuation of MBP staining (demyelination) and moderate‐to‐extensive gliosis associated with increased size of intervening spaces. Inflammatory cell lesions, however, were notably absent. Biophysical analysis of isolated spinal cord myelin from affected rats demonstrated a distorted distribution in subfraction densities and the appearance of extra‐myelin proteins in the light myelin subfraction. Immunologically, chronically affected animals were unresponsive to the encephalitogenic determinant on GP‐BP, although other BP determinants elicited strong delayed type hypersensitivity (DTH) reactions in rats immunized initially with GP‐BP in CFA. These data show that ongoing neurologic dysfunction can be induced in the Lewis rat by a GP‐BP specific T‐lymphocyte line; they suggest that unremitting clinical signs can persist in the absence both of inflammatory lesions in the CNS and of pronounced immunologic responsiveness to the encephalitogenic determinant of GP‐BP.
Uptake of GABA and nipecotic acid in astrocytes and neurons in primary cultures: Changes in the sodium coupling ratio during differentiation
Authors: Larsson O.M. , Hertz L. , Schousboe A.
Published: January 2005

The coupling ratio betwen sodium and GABA or the GABA analog, (RS)‐nipecotic acid, in neuronal and glial uptake of GABA or nipecotic acid was investigated as a function of the morphological differentiation of these cell types in primary cultures. Both in neurons and astrocytes a high‐affinity uptake of GABA and nipecotic acid was observed ragardless of the degree of differentiation. The values of Km were essentially identical in undifferentiated and differentiated cells. On the other hand, Vmax was significantly increased both in 10‐day‐old neurons compared to 1‐ and 3‐day‐old neurons and in astrocytes treated with dBcAMP compared to untreated cells, i.e., Vmax increased as a function of differentiation in both cell types. In neurons as well as in astrocytes the morphological differentation resulted in an alteration in the coupling ratio between sodium and GABA. From calculated Hill coefficients it could be deduced that the coupling ratio between sodium and GABA was changed from 1 to 2 during differentiation. Similar results were obtained for (RS)‐nipecotic acid.
Identification of a glycine‐like fragment on the strychnine molecule
Authors: Aprison M. H. , Lipkowitz K. B. , Simon J. R.
Published: January 2005

Strychnine, a complex molecule, antagonizes in some unknown manner the action of glycine, an important inhibitory neurotransmitter in the spinal cord and brainstem of many vertebrates. To help understand the mechanism of this antagonism, we have employed modern computational methods to assess the similarities between these seemingly different molecules. An exhaustive comparison of topological and electronic features of both molecules was made. We have successfully located a glycine‐like fragment in the strychnine molecule that, when compared to glycine, exhibits both topological and electronic charge congruence. The successful location of this glycine‐like fragment allows us to speculate how the large strychnine molecule assumes its role as an antagonist against the inhibitory action of glycine, the simplest amino acid.
Interleukin‐1‐like activity in rat brain: Sources, targets, and effects of injury
Authors: Nieto‐Sampedro M. , Berman M. A.
Published: January 2005

Extracts of injured rat brain contained molecules that shared biological and physicochemical properties with interleukin‐1 (IL‐1). Brain IL‐1‐like activity increased following brain injury in parallel with the increase in astrocyte and fibroblast mitogenic activity. After 3 days postlesion, it reached about 20 times the basal (noninjured) level. Monoclonal antibodies to human IL‐1 inhibited this brain IL‐1‐like activity.

One of the cellular sources of brain IL‐1‐like activity seems to be astroglial cells. Primary cultures of purified rat brain astrocytes secreted into the culture medium more IL‐1 activity than comparable numbers of peripheral blood monocytes. Brain IL‐1, as well as authentic monocyte IL‐1, appear to act on brain glial cells, promoting thymidine incorporation into purified astrocytes in culture.
Investigations on myelinogenesis in vitro: Developmental expression of myelin basic protein mRNA and its regulation by thyroid hormone in primary cerebral cell cultures from embryonic mice
Authors: Shanker G. , Campagnoni A. T. , Pieringer R. A.
Published: January 2005

The concentration of myelin basic protein (MBP) mRNA in primary cultures of cells dissociated from embryonic mouse cerebra and grown in the presence of varying amounts of thyroid hormone was measured using a 32P‐labeled cDNA probe and a dot‐blot procedure. The cDNA probe contained 1.85 kilobases of the gene for MBP. The concentration of mRNA specific for MBP in control cells grown on a medium containing normal (euthyroid) calf serum increased with increasing age of culture. The greatest increase occurred between 15 and 35 days in culture (5.25‐fold increase); whereas between 35 and 50 days in culture, the rate of accumulation slowed to yield a net increase of MBP mRNA of only 10%. The quantity of MBP mRNA was drastically diminished at all ages studied when the cells were grown from the sixth day onward on a medium containing hypothyroid calf serum. Although the amount of MBP mRNA in hypothyroidtreated cells did increase, the change in concentration was less (3.43‐fold), and it peaked earlier (at 30 days). Unlike the euthyroid cells, after 30 days the MBP mRNA actually fell in the hypothyroid‐treated cells. If hypothyroid media were supplemented with triiodothyronine (T3) on the eighth day in culture, the quantity of MBP mRNA in the cells was restored almost completely to the levels found in the control euthyroid cells at all ages. Therefore, the regulation of the synthesis of MBP by thyroid hormone is at least in part a pretranslational event; that is, thyroid hormone adjusts the concentration of the mRNA specific for MBP.
Transplant‐derived astrocytes migrate into host lumbar and cervical spinal cord after implantation of El4 fetal cerebral cortex into adult thoracic spinal cord
Authors: Goldberg W. J. , Bernstein J. J.
Published: January 2005

Fourteen‐day gestation fetal cerebral cortex homografts were transplanted into the thoracic (T6) spinal cord between the left dorsal column and dorsal horn of adult host rats. The transplants were soaked in 2.0 pg/ml of the lectin Phaseolus vulgaris leucoagglutinin (PHAL) prior to implantation. Transplanted host spinal cords were utilized at 7, 14, and 24 d and at 1 and 2 months postimplantation. Paraffin‐sectioned spinal cords were double labeled for PHAL and glial fibrillary acidic protein (GFAP) by using FITC‐ and RITC‐conjugated secondary antisera, respectively. Montages of FITC‐ and RITC‐positive cells were analyzed for cells containing both fluorescences. Double‐labeled cells (PHAL‐GFAP) were transplant‐derived astrocytes. Transplant‐derived astrocytes were observed to initiate migration in the white matter columns of the host at approximately 14 d after transplantation. Double‐labeled astrocytes were observed in cervical and lumbar spinal cord of the host (ca. 3.5 cm away from the center of the transplant) at 2 months postoperative. These astrocytes migrated at approximately 0.76 mm a day (after a 14‐d delay). At 2 months, transplant‐derived astrocytes composed as much as 50% of the astrocytes in the white matter of the host 2.0 mm from the transplant. The migrated astrocytes were hypertrophied and appeared reactive. Astrocytes in spinal gray matter only migrate about 1.0 mm from the graft‐host interface. Transplant‐derived astrocytes can migrate the entire length of the spinal cord white matter.
Conditioned enhancement of natural killer cell activity, but not interferon, with camphor or saccharin‐Licl conditioned stimulus
Authors: Ghanta V. K. , Hiramoto N. S. , Solvason H. B. , Tyring S. K. , Spector N. H. , Hiramoto R. N.
Published: January 2005

Pavlovian conditioning of the natural killer (NK) cell response has been demonstrated by pairing camphor with polyinosinic: polycytidylic acid (poly I: C) in nine association trials. The NK cell response could be conditioned also by using combined saccharin and lithium chlorid (LiCl) as the conditioned stimulus. The camphor and saccharin‐LiCl paradigms were tested to determine if the conditioned NK cell activity was the result of conditioning of the interferon response. Interferon levels were measured at 6 hr and NK cell activity at 24 hr after application of the conditioned stimulus. The interferon levels measured in separate experiments were not uniformly elevated in conditioned animals compared with controls.
Preproenkephalin mRNA in T‐Cells, macrophages, and mast cells
Authors: Martin J. , Prystowsky M. B. , Angeletti R. H.
Published: January 2005

The presence of preproenkephaline mRNA in tumor cell lines derived from myeloid and mast cells was analyzed by RNA dot blot hybridization. A B‐cell lymphoma was negative for preproenkephalin mRNA, but several T‐cell lymphomas were positive. A mastocytoma and two macrophage cell lines were found to have high levels of preproenkephalin mRNA. Purified natural macrophages and mast cells also possessed easily detectable levels of this mRNA.
Novel processing pathway for proopiomelanocortin in lymphocytes: Endotoxin induction of a new prohormone‐Cleaving enzyme
Authors: Harbour D. V. , Smith E. M. , Blalock J. E.
Published: January 2005

We have discovered that the immune system processes proopiomelanocortin (POMC) products differently depending on the stimulus for induction. We have shown that corticotropin‐releasing factor (CRF) induces the lymphocytes from C3HeB/FeJ lipopolysaccharide (LPS)‐sensitive mice to produce adrenocorticotropin (ACTH) 1‐39 and β‐endorphin, whereas LPS induces these lymphocytes to produce ACTH 1‐23 to 26 and α‐ or γ‐endorphin. We have proposed that the smaller species of ACTH and endorphin are proteolytic cleavage products from ACTH 1‐39 and β‐endorphin analysis of C3HeB/FeJ LPS‐treatment B lymphocyte lysates showed an enzymatic activity at pH 5 but not pH 7 that cleaved ACTH 1‐39 into a smaller ACTH 1‐23 to 26. The B lymphocytes from C3HeB/FeJ (LPS‐resistant) mice expressed but did not process proopiomelanocortin after LPS or CRF treatment, nor did their B cells express the aforementioned enzymatic activity. Taken together, these data suggest a unique processing pathway in LPS‐treated B lymphocytes and one in which immunoreactive (ir)‐endorphins may play a role in the pathophysiology of endotoxic shock.
Neuroendocrine and metabolic responses induced by Interleukin‐1
Authors: Besedovsky H. , del Rey A.
Published: January 2005

We have previously demonstrated that Interleukin‐1 (IL‐1), a cytokine mainly produced by activated monocytes, stimulates the pituitary–adrenal axis and affects glucose homeostasis. Comparative studies revealed that a β form of recombinant human IL‐1, similar to the mature peptide secreted naturally, is more powerful than other preparations of this monokine in stimulating adrenocorticotrophic hormone (ACTH) and corticosteone output. Another monokine, tumor necrosis factor (TNF), does not share with IL‐1 the capacity to induce such effects. In extending out studies to rats, we showed that increased ACTH and blood corticosterone levels are also induced by IL‐1 in this species. Another in vivo activity of IL‐1 relates to its capacity to induce a reduction in blood glucose levels. Our studies strongly suggest that, as opposed to other effects elicited by IL‐1, blockade of prostaglandin synthesis does not effect the capacity of IL‐1 to stimulate insulin output and produce hypoglycemia. Administration of IL‐1 to adrenalectomized mice, which are defective in counterregulatory mechanisms of glucose homeostasis, resulted in marked hypoglycemia. In contrast to the response observed in normal mice, in adrenalectomized animals this effect was paralleled by decreased blood insulin levels. IL‐1 was also injected into alloxan‐diabetic mice. A marked reduction in blood glucose levels occurred in these animals. This effect was already noticeable 1 hr after injection. After 2 hr and for at least another 6 hr, glucose levels of alloxan‐treated mice injected with IL‐1 remained within the normal range. A possible early increase in release of residual insulin in diabetic animals was ruled out. These data, together with previous studies, suggest that IL‐1‐induced hypoglycemia is not only mediated by its insulin secretagogue effect. The fact that IL‐1, a key mediator of immunological and inflammatory reponses, affects crucial mechanisms under control of the central nervous system suggests that it play a major role in neuroendocrine immunoregulation.
Glial fibrillary acidic protein in chronic relapsing experimental allergic encephalomyelitis in SJL/J mice
Authors: Smith M. E. , Eng L. F.
Published: January 2005

The gliotic scar in the demyelinated plaque is a prominent feature of the pathology of multiple sclerosis. Chronic relapsing experimental allergic encephalomyelitis (EAE) in the SJL/J mouse has many characteristics in common with multiple sclerosis in the human, including the development of intense gliosis during the course of the demyelinating disease. With the use of antibody to the astrocyte marker glial fibrillary acidic protein (GFAP) we have measured the increase of GFAP over an 11‐month course of chronic EAE. Intense staining of astrocyte fibers was seen around EAE lesions, which were most frequently observed in t he cerebellum, periventricular areas, and in the spinal cord, The relative amount of GFAP was estimated by preparation of cytoskeletal proteins from the affected CNS areas, separation of proteins by polyacrylamide gel electrophoresis, and quantitation of GFAP in relation to the 70‐kD neurofilament protein (NF) in gel scans. The ratio GFAP/70‐kD NF protein in control animals did not change significantly over 11 months, whereas this ratio gradually increased to 2.54 in animals with chronic relapsing EAE 6 months after immunization. Although some decrease of neural fibers may have contributed partially to this change in ratio, the amounts of GFAP were greatly increased.

These results indicate that the SJL/J mouse with chronic relapsing EAE provides an excellent model with which to investigate the formation and development of the gliotic plaque analogous to that seen in demyelinated areas in multiple sclerosis tissue.
HLA‐DR restriction‐fragment‐length polymorphisms in narcolepsy
Authors: Holloman J. D. , Bell J. I. , Kilduff T. S. , Dement W. C. , Guilleminault C. , McDevitt H. O.
Published: January 2005

Seventeen white patients from the Stanford Sleep Disorders Clinic compaining of excessive daytime somnolence (EDS) were selected for restriction‐fragment‐length polymorphism (RFLP) studies. Fourteen of the patients with clinically diagnosed narcolepsy were seropositive for DR2. RFLP analysis of these patients compared with a homozygous DR2‐Dw2 cell line failed to reveal any polymorphism when digested with six restriction endonucleases and hybridized with three different cDNA probes. None of the three patients with central nervous system hypersomnia, a syndrom similar to narcolepsy, were DR2‐positive. We conclude that any polymorphism of the DRβ, DQα or DQβ, genes of DR2 narcoleptics that might distinguish them from DR2 nomals cannot be resolved through RELP analysis.
An in vivo and in vitro analysis of systemic immune function in mice with histologic evidence of neural transplant rejection
Authors: Nicholas M. K. , Stefansson K. , Antel J. P. , Arnason B. G. W.
Published: January 2005

Histologic and immunocytochemical analyses of fetal neocortical tissue transplanted to the lateral vnetricle of ibred adult mice indicate that this tissue survives transplantation well if the donor and host are isogeneic. The major histocompatibility complex (MHC) of the mouse is known as the H‐2 locus. H‐2‐incompatible neural transplants (allografts), unlike their H‐2‐identical counterpart (isografts), are characterized by the presence of T cells comprising both major T‐cell subsets and macrophages, and by a marked increase in the expression of both class I and class II (Ia) MHC antigens. These findings suggest a recognition of H‐2 alloantigens by the host's immune system followed by an appropriate effector response. We report here out attempts to demonstrate systemic host sensitization to alloantigens in micebearing H‐2‐incompatible intraventricular neural transplants. We measured the time to rejection of orthotopic skin grafts subsequent neural transplantation, splenocyte proliferative responses to alloantigens in mixed lymphocyte cultures (MLC), and class I‐restricted antigen‐specific cytolytic T lymphocyte (CTL) activity. No significant differences were found in any of these tests of host systemic sensitization between mice with allogeneic neural transplants and those with isogeneic transplants or control animals. We conclude that intraventricular neural transplants, while recognized and affected by cells of the host's immune system, do not elicit a detectable systemic sensitization to class I H‐2 alloantigens. Rejection of neural transplants may depend on sensitization to class II H‐2 alloantigens, to so‐called minor histocompatibility antigens, or some combination thereof.
Receptor‐mediated transcytosis of transferrin across the blood‐brain barrier
Authors: Fishman J. B. , Rubin J. B. , Handrahan J. V. , Connor J. R. , Fine R. E.
Published: January 2005

The perfusion of rat brain with 125I‐transferrin resulted in a receptor‐mediated uptake of transferrin into the endothelium of the blood‐brain barrier followed by its detecation in the brain. During a pulsechase experiment, 125I‐transferrin accumulated in the endothelial cells during the pulse, with a decrease of this intraendothelial radioactivity during the chase associated a concomitant increase in the nonvascular elements of the brain. The receptor‐mediated movement of transferrin across the blood‐brain barrier suggests that the brain may derive its iron through the transcytosis of iron‐loaded transferrin across the brain microvasculature. We discuss the likelihood that aluminum and other potentially toxic heavy metals, which also bind tightly to transferrin, may enter the brain by this pathway. We also discuss the possibility that other large molecules including neuroactive peptides and neurotrophic viruses may enter the brain through a similar receptor‐mediated, vesicular transcytotic route.
Preferential adhesion of brain astrocytes to laminin and central neurites to astrocytes
Authors: Pixley S. K. R. , Nieto‐Sampedro M. , Cotman C. W.
Published: January 2005

Astrocytes from either fetal or newborn rat brain adhered preferentially to surfaces coated with active laminin. Neurites from septal neurons did not show a preference for active over inactive laminin. However, when given a choice between laminin and an astrocytic surface, septal cells preferred to extend neurities over astroglial processes. Hence, laminin paths can guide astrocyte migration, which, in turn, can guide neurite elongation.
Differentiation of cerebellar bipotential glial preecurosrs into oligodendrocytes in primary culture: Development profile of surface antigens and mitotic activity
Authors: Levi G. , Aloisi F. , Wilkin G. P.
Published: January 2005

We have analyzed the changes in surface antigenic properties of cerebellar bipotential precursors of oligodendrocytes and type‐2 astrocytes during their differention into oligodendrocytes in serum‐free cultures and relationship between antigen expression and proliferation of these cells. Double immunofluorescence experiments with different monoclonal fluroescence experiments with different monoclonal antibodies (mabs) performed at various stages in vitro and immunocytolysis experiments provided evidence for the following antigenic development profile: at early stages in culture the progenitor cells are recognized by the mabs A2B5 and LB1 (which bind to surface gangliosides) but not by other mabs known to label immature or mature oligodendrocytes (04, 01, and anti‐galactocerebroside [GalC]). A few days later, the precursors start to express the 04 antigen; at this stage they maintain a bipotential nature and, in the presence of serum, they differentiate into type‐2 astrocytes. If maintained in serum‐free medium, the progenitor cells enter the oligodendrocyte differentiation compartment, acquiring GalC postivity. Soon after compartment GalC+, the cells lose both bipotentiality and the surface antigens binding A2B5 and LB1. They conserve, however, the antigen binding 04. Experiments of [3H]thymidine autoradiography combined with immunofluorescence showed that a greater proportion of the LB1+ cells incroporated the radioactive nucleoside into their nuclei as compared to the 04+ cells. No incorporation was present in GalC+ oligodendrocytes.