bluuebunny/crossref_metadata_2025 huggingface dataset:
The Remarkable Structure of Lithium Cyanide/Isocyanide
Authors: Von Ragué Schleyer Paul , Sawaryn Andrzej , Reed Alan E. , Hobza Pavel
Published: January 2005

Electron correlation corrections have a considerable influence on the relative stabilities of lithium isocyanide (1), lithium cyanide (2), and the bridged form, 3. While Hartree‐Fock theory finds 1 to be most stable and 3 not to be a minimum, MP2/6‐31G* optimization indicates 3 to be the global minimum. At higher levels employing full fourth‐order Møller‐Plesset theory and a quadruply split valence and polarized basis set (MP4STDQ/6‐311+G*), 2 is only about 2 kcal/mol less stable than 1 and 3, which are indicated to have nearly the same energy. LiNC thus is similar to C(Na)N and C(K)N, both of which are known to prefer T‐shaped (bridged) structures in the gas phase. However, to an even greater extent than formerly realized, rotation of the lithium cation around the cyanide anion nucleus should be practically free. ΔH (LiCN) = 32.8 kcal/mol is estimated from the calculated lithium cation affinity of 151.2 kcal/mol. In addition, we find at the MP4SDTQ/6‐31+G*//MP2/6‐31G* level that the bridged form of NaCN is favored by 2–3 kcal/mol over the corresponding linear forms, which have nearly the same energy.
Free energy perturbation calculations on models of active sites: Applications to adenosine deaminase inhibitors
Authors: Hansen Lillian M. , Kollman Peter A.
Published: January 2005

Free energy perturbation calculations were performed to determine the free energy of binding associated with the presence of perhaps an unusual hydroxyl group in the transition state analog of nebularine, an inhibitor of the enzyme adenosine deaminase. The presence of a single hydroxyl group in this inhibitor has been found to contribute −9.8 kcal/mol to the free energy of binding, with a 108‐fold increase in the binding affinity by the enzyme. In this work, we calculate the difference in solvation free energy for the 1,6‐dihydropurine complex versus that of the 6‐hydroxyl‐1,6‐dihydropurine complex to determine if this marked increase in binding affinity is attributed to an unusually hydrophobic hydroxyl group. The calculated ΔG associated for the solvation free energy is −11.8 kcal/mol. This large change in the solvation free energy suggests that this hydroxyl is instead unusually hydrophilic and that the difference in free energy of interaction for the two inhibitors to the enzyme must be at least ca. 20 kcal/mol. Although the crystal structure for adenosine deaminase is currently not known, we attempt to mimic the nature of the active site by constructing models which simulate the enzyme‐inhibitor complex. We present a first attempt at determining the change in free energy of binding for a system in which structural data for the enzyme is incomplete. To do this, we construct what we believe is a minimal model of the binding between adenosine deaminase and an inhibitor. The active site is simulated as a single charged carboxyl group which can form a hydrogen bond with the hydroxyl group of the analog. Two different carboxyl anion models are used. In the first model, the association is modeled between an acetic acid anion and the modified inhibitor. The second model consists of a hydrophobic amino acid pocket with an interior Glu residue in the active site. From these models we calculate the change in free energy of association and the overall change in free energy of binding. We calculate the free energies of interaction both in the absence and presence of water. We conclude from this that the presence of a single suitably placed‐CO−2 group probably cannot explain the binding effect of the‐OH group and that additional interactions will be found in the adenosine deaminase active site.
A hybrid compartmental model for the alligator Purkinje cell. I: Preferred somatopetal conduction of dendritic spikes and soma‐axon interaction
Authors: Pottala Erik W. , Mortimer James A.
Published: January 2005

A compartmental hardware model of an alligator Purkinje cell is described, consisting of a branched dendritic tree with four zones of spike generation and electrically excitable soma and initial‐segment regions. Passive properties of the model compartments are represented by a cable analog circuit. Simulated action potentials, generated by a combination of depolarizing and hyperpolarizing conductance changes, are triggered in active compartments when the simulated membrane potential passes through preset thresholds. These were set at values corresponding to 28mV depolarization in the dendrites, 22 mV in the soma, and 7 mV in the initial‐segment compartment. Synaptic inputs consisting of brief (0.35 msec) rectangular conductance changes give rise to exponentially decaying postsynaptic potentials in the input compartment which are electrotonically spread to other compartments. Orthodromic activation of the model neuron by computer‐generated random pulse trains generates a simple discharge in the initial‐segment compartment without evoking complex spikes. Synchronized excitatory input to the same compartments, however, does evoke a complex spike response in the soma and initial segment, coupled with dendritic spikes, Following antidromic activation of the model neuron, dendritic spikes are not generated, demonstrating a tendency in the dendritic tree for preferential conduction of spikes toward the soma. Investigation of some of the factors underlying this tendency suggests that variations in voltage attenuation due to dendritic geometry, convergence of electrotonically spread dendritic spikes, and the relative durations of dendritic and somatic action potentials may contribute to it. The presence of a threshold gradient in the dendritic tree, proposed by Llinás and his coworkers, was not found to be necessary to explain this tendency toward somatopetal conduction, although it cannot be excluded by the model. Examination of the role of the conically shaped initial‐segment region suggests that this zone may provide a low‐pass filter for signals conducted electrotonically from the axon to the soma, blocking repolarization of the soma during the complex spike burst generated in the axon.
An adrenergic mechanism for visual neuroregulation
Authors: Coppola Robert J. , Rolek Dennis F. , Kramer Stanley Z.
Published: January 2005

Studies of the diffuse electroretinogram (ERG) and evoked responses in the optic lobe in the baby chick have shown that epinephrine produces changes in both. Epinephrine produces enhancement of the a‐wave and depression of the b‐ and d‐waves in both intact and isolated eyes. In the optic lobe it produces enhancement of both components of the evoked response. The presence of epinephrine in the vertebrate retina and the optic lobe of chicks suggests that these changes may reflect the involvement of epinephrine in synaptic transmission.
Spontaneous and flash‐induced multiple unit activity in the hypothalamic gonadotropic areas: Effects of photoperiod and of optic nerve section in quail
Authors: Oliver J. , Baylé J. D.
Published: January 2005

The present study was devoted to recording of spontaneous and flash‐evoked multiple unit discharges in the gonadotropic areas of the quail hypothalamus and to investigate the influence of the length of the day and of bilateral optic nerve section.

Blinding resulted in a marked decrease of MUA in the tuberal region which was much less important in the dorsal part of the nuclear infundibular complex. Burst and continuous patterns of discharges subsisted in tuberal and dorsal areas of blinded quail but responses to flash were completely suppressed.

Lengthening the photofraction led to a very strong reduction of firing rates in both tuberal and dorsal hypothalamic regions. Furthermore, flash stimulation was no longer effective in intact photostimulated quail (18L‐6D).

Flash‐evoked modification of MUA in gonadotropic regions were differentiated in a clear‐cut fashion from specific responses which were obtained in the visual system.
Cerebellar contributions to the papez circuit
Authors: Snider Ray S. , Maiti A.
Published: January 2005

Cerebellar influences on the various substructures in the Papez Circuit are indicated by the following.

Anatomical studies indicate that the major midbrain areas to which this circuit projects are : (1) ventral tegmental area; (2) interpeduncular area; and (3) periaquenductal gray areas; and these same areas project back to the limbic system. There are projections to these regions from the cerebellar nuclei, as indicated by terminal degeneration studies which show that cerebellar nuclei connect, mostly by fine fibers, with a continuum of cells located on either side of the midline in the ventral tegmentum of the midbrain. Observations that the cerebellum also projects to the locus ceruleus (NA system) and VTA (DA system) indicate that cerebellar influences can also reach the limbic areas via the catecholamine fiber bundles.

Electrophysiological studies indicate that vermian and fastigial stimulation induce evoked responses in the basolateral amygdala, the hippocampus, and the septum, with laterncies to the peak of first wave ranging from 4 to 8 msec and to the second wave of 16–29 msec. Citations from the physiological literature indicate that electrical stimulation of the cerebellum, especially the vermis, can modify a wide range of responses which involve fuctional activities of either the sympathetic or parasympathetic nervous systems.

Studies on electrically induced afterdischarges in the septum, Hippocampus, and amygdala indicated that cerebellar stimulation can shorten the duration of or terminate that afterdischarges, and the site of lowest threshold is the midline cortex. Focal cooling of the vermis promotes prolongation of the afterdischarges as does preteatment of animals with 6‐OH dopamine. Chemical lesions in the catecholamine system induced by 6‐OH dopamine reduce the effectiveness of the cerebellar stimulation, as do lesions of nucleus fastigii, These data are interpreted to indicate that the cerebellum can exert a tonic suppressor (inhibitory?) influence on substructures within the Papez Cricuit.

Citations from animal behavioral studies indicate that electrical stimulation of the anterior cerebellum can induce responses such as arousal, predatory attack, and feeding which mimic those obtained by amygdaloid stimulation. Fastigial stimulation can produce drowsiness and EEG changes which resemble the sleep patterns resulting from stimulation of the ventral amygdala.
Cellular and subcellular distribution of the S‐100 protein in rabbit and rat central nervous system
Authors: Haglid K. G. , Hamberger A. , Hansson H.‐A. , Hydén H. , Persson L. , Rönnbäck L.
Published: January 2005

The cellular and subcellular distribution of the S‐100 protein in rabbit and rat central nervous system was studied both quantitatively and qualitatively. Microcomplement fixation estimations on bulk‐prepared neuronal and glial cells showed at least five to six times higher amounts of water‐soluble S‐100 protein. High levels of S‐100 protein were found in mitochondria and soluble protein fractions. Immunoelectron microscope investigations demonstrated S‐100 protein in neuronal structures, such as the postsynaptic membrane and part of the plasma membrane. Among glial cells the astrocytic filaments contained high levels of S‐100 protein. S‐100 was also found in most subcellular membranes of astrocytes and oligodendrocytes.
Effects of cannabis sativa on ultrastructure of the synapse in monkey brain
Authors: Harper Jon W. , Heath Robert G. , Myers William A.
Published: January 2005

Electron microscopic study of brain tissues of monkeys chronically exposed to psychoative Cannabis showed widening of the synaptic cleft, electron opaque material in the cleft and in pre‐ and postsynaptic regions, and some “clumping”: of synaptic vesicles. In contrast, tissues of control monkeys showed no ultrastructural changes.
Pharmacology of the brachium conjunctivum: Red nucleus synaptic system in the baboon
Authors: Huffman Ronald D. , Davis Ross
Published: January 2005

Acetylcholine, biogenic amines, and certain amino acids were applied by microintophoresis to parvicellular and magnocellular red nucleus (RN) neurons of baboon while recording brachium conjunctivum (BC)‐evoked and amino acid‐evoked unit discharge from these neurons. Glycine, γ‐aminobutyric acid, and β‐alanine were potent depressants of BC‐RN synaptic transmission, amino acid‐evoked firing, and spontaneous activity of all RN neurons studied. Glycine was clearly more potent than the other 2 depressant amino acids. L‐Glutamic and DL‐homocysteic acid were strong excitants of all RN neurons tested. Dopamine, noradrenaline, and 5‐hydroxytryptamine depressed the excitability of both parvicellular and magnocellular RN neurons; no excitatory effects were observed with these biogenic amines on RN neurons. Acetylcholine increased the rate of firing of spontaneously discharging parvicellular RN neurons and facilitated the amino acid‐induced firing of these same neurons. Acetylcholine did not facilitate BC‐RN synaptic transmission nor could this transmission be blocked by cholinergic antagonists. Unlike parvicellular RN neurons, the responsiveness of magnocellular neurons was either unaltered by acetylcholine or slightly decereased. These experiments demonstrate a difference in the pharmacologic responsiveness of parvicellular and magnocellular RN neurons to acetylcholine but do not provide evidence for a cholinergic input to RN via the brachium conjunctivum.
The effect of chronic administration of d‐amphetamine on regional changes in catecholamines in the rat brain
Authors: Lynch Marina , Kenny Marie , Leonard B. E.
Published: January 2005

Changes in the dopamine, noradrenaline and dihydroxyphenylacetic acid (DOPAC) concentrations were determined in 6 regions of the rat brain following chronic amphetamine administration for 2 weeks and at intervals of up to 7 days after drug withdrawal. Chronic amphetamine administration caused pronounced stereotypy; following withdrawal the animals were behaviourally depressed for at least 7 days. The noradrenaline concentration of all brain areas investigated was decreased throughout most of the experimental period; only in the midbrain and the olfactory lobes did the noradrenaline concentration return to control values 7 days after drug withdrawal. In contrast, the dopamine concentration was raised in the midbrain, hippocampus, and olfactory lobes following drug administration but reduced in the striatum, brain stem, and amygdaloid cortex. Following amphetamine withdrawal, most of the brain areas showed a decrease in the dopamine concentration apart from the olfactory lobes which showed an increase and the midbrain and hippocampus which were largely unchanged. The concentrations of the intraneuronal metabolite DOPAC were decreased in most brain areas throughout the duration of the experiment, suggesting that the turnover of dopamine was decreased. These results are discussed with reference to the possible decreased activity in tyrosine hydroxylase both during drug treatment and after withdrawal.
Ethanol tolerance: Evidence of “protective” effects on brains of adult rats
Authors: Klemm W. R.
Published: January 2005

Previous research with two biochemical markers of acute ethanol damage (sialic acid and 2‐deoxyribose) raised the possibility that tolerance developed by chronic ingestion of ethanol. However, this hypothesis required demonstration that chronic consumption was not damaging. This issue was investigated histologically in adult rats that voluntarily consumed massive doses of ethanol daily (range of 11 to 18 gm/kg/rat/day) for 28 days. By all indices (thickness measures of neocortex, hippocampus, and cerebellar molecular layer; and specific cell counts of neocortex and cerebellum), none of the ethanol‐exposed rats, even those with intentional nutritional deficiencies, revealed any physical sign of damage compared to control rats.
An interaction between the pineal gland and olfactory deprivation in potentiating the effects of melatonin on gonads, accessory sex organs, and prolactin in male rats
Authors: Blask David E. , Nodelman Jacqueline L.
Published: January 2005

Male rats (25–26 days of age) housed with 14 hours of light per day (lights on 0600–2000 hours) were either olfactory bulbectomized (rendering them anosmic), bulbectomized plus pinealectomized (Pinx), or left intact. On the day following the operations, intact, anosmic, and anosmic‐Pinx animals began receiving single, daily afternoon (1700–1800 hours) subcutaneous injections of 50 μg of melatonin (MEL) for six weeks, while an additional group of intact controls received injections of diluent. At the end of this period, body, anterior pituitary, testicular, and seminal vesicle weights were significantly reduced in intact‐MEL‐treated animals. Anosmic animals that had been treated with MEL experienced a further, highly significant, 65%, 90%, and 85% depression in testicular, seminal vesicle, and ventral prostate weights, respectively, as compared with intact control and MEL‐treated rats. Additionally, both body and anterior pituitary weights were significantly decreased in MEL‐treated, anosmic rats. Anosmic‐Pinx rats treated with MEL had organ and body weights that were intermediate between those of intact‐MEL and anosmic‐MEL‐treated animals. Pituitary and serum levels of prolactin (Prl) were significantly lower in anosmic‐MEL‐treated rats than in intact or intact‐MEL‐treated groups. Similarly, Prl levels were depressed in the anosmic‐Pinx rats treated with MEL; however, serum Prl was not statistically lower than in intact or intact‐MEL‐treated animals. These results indicate that anosmic male rats have an increased sensitivity to antigonadotrophic and Prl‐inhibitory effects of MEL. Furthermore, the data suggest that the presence of the pineal gland in anosmic rats is important in permitting anosmia maximally to sensitize the neuroendocrine‐reproductive axis to the antigonadotrophic effects of exogenously administered MEL.
Isolation of ribosome containing synaptosome subpopulation with active in vitro protein synthesis
Authors: Verity M. Anthony , Brown W. Jann , Cheung M.
Published: January 2005

Subpopulations of synaptosomes harvested from neonatal rat brain cortices revealed a differential ability to synthesize protein in vitro. Incubation of synaptosomes with radiolbeled leucine, followed by continuous sucrose gradient centrifugation produced an asymmetric shift in the radioactivity toward the higher density sucrose fractions. The bulk of the mitochondrial cytochrome c‐oxidase activity was also found in these fractions, however, subfractionation studies of osmotically‐lysed synaptosomes suggested that the newly‐synthesized proteins reside in an osmotically sensitive, non‐mitochondrial compartment.

The ability of each subpopulation of synaptosomes to synthesize protein in vitro was assessed after their isolation from linear continuous sucrose gradients. There was an enrichment of highly active protein synthesizing particles in the “heavy” subpopulations of neonatal synaptosomes. The inhibitory effects of chloramphenicol and cycloheximide on the protein synthesis in these particles were similar to those of the original synaptosome fraction. Electron microscopic analysis revealed an increase in the numbers of ribosome‐containing structures resembling dendritic dendritic and axonal growth cones.
The uptake of 3‐deoxy‐3‐fluoro‐D‐Glucose by synaptosomes from rat brain cortex
Authors: Halton D. M. , Taylor N. F. , Lopes D. P.
Published: January 2005

D‐[3‐3H]‐3‐deoxy‐3‐fluoroglucose was synthesized chemically and shown to be transported into rat brain synaptosomes by a saturable process with a Km 6.2 × 10−4 M and a Vmax 2.8 nmole · mg protein−1. After an initial, rapid period of transport, further uptake of the fluorosugar is restricted by the rate of its phosphorylation. Both D‐glucose and cytochalasin B are competitive inhibitors of 3‐deoxy‐3‐fluoro‐D‐glucose transport with Ki values of 93 μm and 6.0 × 10−7 M, respectively. Phloretin, N‐ethylmaleimide and p‐chloromercuribenzoate also inhibit the fluorosugar uptake, whereas ouabain and changes in K+, Na+, Mg2+ and Ca2+ ions have only a small effect. The recorded 3‐deoxy‐3‐fluoro‐D‐glucose influx is slightly reduced by potassium cyanide, antimycin A, 2,4‐dinitrophenol, and rotenone. The uptake reduction caused by these four reagents is relieved by the addition of exogenous ATP. The possible influence of hexokinase activity on the uptake process is discussed.
Role of nerve growth factor in ethylnitrosourea‐induced neural carcinogenesis
Authors: Vinores Stanley A. , Perez‐Polo J. Regino
Published: January 2005

The sensitivity of the rat nervous system to malignant transformation by ethylnitrosourea (ENU) is a function of age at treatment. From late gestation, nervous structures decrease in sensitivity with age as non‐neural structures increase in susceptibility. There is a decrease in the proportion of neural tumors induced by ENU and an increase in survival time when nerve growth factor (NGF) levels are elevated in the fetal or neonatal stage. If antibodies directed against mouse beta‐NGF (anti‐NGF) are administered prior to neonatal ENU treatment, neural tumors appear earlier, although in the same proportion as with treatment by ENU alone. This effect is not observed if the ENU is administered first. This phenomenon seems to be attributed to an increased number of trigeminal nerve neurinomas, which have a shorter latent period than other nervous system tumors. The induced neurological tumors in rats treated neonatally with anti‐NGF prior to ENU seem to be almost exclusively neurinomas in the peripheral nervous system. Fetal anti‐NGF treatment leads to an increased number of intracerebral gliomas and a longer survival time, which corresponds to the longer latent period of these tumors. The role of NGF in the sensitivity of the rat nervous system to carcinogenesis by ENU and its possible implications in the development of the nervous system are discussed.
Respiration in rat cerebral astrocytes from primary culture
Authors: Olson James E. , Holtzman David
Published: January 2005

Respiration was measured polarographically in astrocytes from dissociated neonatal rat cerebra grown in primary culture. Cells grown in a modified Eagle's minimal essential medium containing 20% fetal calf serum for 14 to 21 days respired at a rate of 34.23 n at. equiv 0 mg protein−1 min−1. Cells which were grown in this medium for 14 days and then grown in serum‐free medium containing 0.25 mM dibutyrylcyclic‐AMP (dbcAMP) for three to six days, had respiratory rates 20% higher than in cells from untreated control cultures (P = 0.005). Maximal inhibition by oligomycin required 200 p mole per mg protein for dbcAMP‐treated cells and 80 pmole per mg protein for untreated cells. Oligomycin inhibited the respiration of both dbcAMP‐treated and untreated cells by about 70%. At a concentration of 175 nmole per mg protein, 2,4‐dinitrophenol (DNP) produced a maximal stimulation of respiration, which was 191% of the resting rate in dbc AMP‐treated cells and 218% of the resting rate in untreated cells. The maximal DNP‐stimulated respiratory rates were the same in dbcAMP‐treated and untreated cells.
Metabolic neural mapping in neonatal rats
Authors: DiRocco Richard J. , Hall W. G.
Published: January 2005

Functional neural mapping by 14C‐deoxyglucose autoradiography in adult rats has shown that increases in neural metabolic rate that are coupled to increased neurophysiological activity are more evident in axon terminals and dendrites than neuron cell bodies. Regions containing architectonically well‐defined concentrations of terminals and dendrites (neuropil) have high metabolic rates when the neuropil is physiologically active. In neonatal rats, however, we find that regions containing well‐defined groupings of neuron cell bodies have high metabolic rates in 14C‐deoxyglucose autoradiograms. The striking difference between the morphological appearance of 14C‐deoxyglucose autoradiograms obtained from neonatal and adult rats is probably related to developmental changes in morphometric features of differentiating neurons, as well as associated changes in type and locus of neural work performed.
Glucocorticoid receptor properties and glucocorticoid regulation of glutamine synthetase activity in sensitive C6 and resistant C6H glial cells
Authors: Holbrook Nikki J. , Grasso Robert J. , Hackney John F.
Published: January 2005

The relationship between induction of glutamine synthetase activity by dexamethasone and binding of the steroid to cytosolic glucocorticoid receptors was examined in sensitive C6 and resistant C6H glial cell cultures.

Glutamine synthetase activity increased 3–4‐fold when C6 cultures were exposed to 7.6 × 10−6 M dexamethasone. This inductive response was reversible, dose‐dependent (ED50 ∼ 2 × 10−8 M), required de novo protein and RNA synthesis, and was elicited only by glucocorticoid steroids. Progesterone, but not epicortisol, antagonized the dexamethasone‐induced enzyme increase. In contrast, only a slight inductive effect was observed in dexamethasone‐treated C6H cells.

Competitive binding assays demonstrated that specific binding of [3H]‐dexa‐methasone to cytosolic receptors was also dose‐dependent. The ED50 was ∼ 10−8 M for both C6 and C6H cells. Scatchard analysis revealed that each C6 cell contained ∼10,800 receptor sites and that the equilibrium dissociation constant (Kd) was 4.5 × 10−9 M. Each C6H cell possessed ∼ 12,200 sites, and the Kd was 6.7 × 10−9 M. Unlabeled dexamethasone and cortisol (but not epicortisol) competed effectively with [3H]‐dexamethasone for binding to cytosolic receptor sites and nuclear sites of both cell types.

These results suggest that induction of glutamine synthetase activity in dexamethasone‐treated C6 cells is a glucocorticoid‐directed response. Since C6H cells are refractory in this regard but contain functional cytosolic receptors which interact with cell nuclei, the basis for their resistance appears to involve some step beyond these cellular processes.
Induction of neurite outgrowth in the IMR‐32 human neuroblastoma cell line by nerve growth factor
Authors: Reynolds C. P. , Perez‐Polo J. R.
Published: January 2005

The nerve growth factor protein (NGF) stimulates neurite outgrowth from embryonic sensory ganglia and sympathetic ganglia at all stages of development. In addition, NGF is required for the maintenance of the differentiated state in adult sympathetic ganglia. A clonal cell line, IMR‐32, derived from a human neuroblastoma was found to contain a population of cells that respond to NGF by exhibiting morphological differentiation. The effect of NGF on these cells is compared with that of other agents known to induce differentiation of IMR‐32, including glioma‐conditioned media.
Bioassay detection of mouse nerve growth factor (mNGF) in the brain of adult mice
Authors: Scott S. M. , Tarris R. , Eveleth D. , Mansfield H. , Weichsel M. E. , Fisher D. A.
Published: January 2005

Two pools of seven brains each from adult Swiss‐Webster mice were homogenized, and supernatants were collected for bioassay. PC‐12 cells were placed in a bioassay plate at time zero, at a concentration of 104 cells per well, and primed for 48 hours in a medium containing 50 ng/ml of mNGF. The PC‐12 cell bioassay for neurite outgrowth was conducted after primed cells were exposed to an NGF‐free medium for 24 hours. Suitable controls for serum toxicity and cell viability were established. The sensitivity of the bioassay approximates 100 pg NGF/ml. The results showed 80–100% neurite outgrowth in wells exposed to brain pool supernatant (BPS) alone, and control level outgrowth (3–8%) in wells containing BPS and specific anti‐β‐NGF antibody. Therefore, the brains of Swiss‐Webster adult mice contain an NGF‐like substance which promotes neurite outgrowth in PC‐12 cells. The substance probably is NGF itself, since the effect is blocked by specific NGF antiserum.
Acute and chronic effects of pentobarbital in relation to postsynaptic GABA receptors: A study with muscimol
Authors: Sivam S. P. , Nabeshima T. , Ho I. K.
Published: January 2005

Muscimol, a GABA agonist, enhanced pentobarbital sleeping time in a dose‐dependent manner. The GABA antagonists such as bicuculline and picrotoxin, and the CNS stimulant such as pentylenetetrazol, inhibited pentobarbital sleeping time; however, all except picrotoxin produced less than 35% maximum inhibition. Picrotoxin, an agent which blocks the chloride ionophore of GABA‐receptor complex, exhibited a parallel dose‐response curve with respect to muscimol. Chronic administration of pentobarbital by pellet implantation induced tolerance as evidenced by decreased sleeping time; the tolerance receded gradually upon abrupt withdrawal. Muscimol enhanced pentobarbital sleeping time both in tolerant and withdrawal mice. Na+‐independent GABA‐receptor binding, using [3H]muscimol as a ligand, was increased after acute and chronic pentobarbital administration; withdrawal of the pentobarbital reversed the increase in receptor population. None of the treatments altered the affinity of [3H]muscimol binding. These results support the contention that pentobarbital (a) directly acts on the postsynaptic chloride ionophore and (b) augments GABA‐mediated postsynaptic effects. The functional significance of the increase in GABA receptor population after pentobarbital treatment is unclear.
A proposed method for the determination of cerebral regional intermediary glucose metabolism in humans in vivo using specifically labeled 11C‐glucose and positron emission transverse tomography (PETT). I. An animal model with 14C‐glucose and rat brain autoradiography
Authors: Sacks William , Sacks Shirley , Badalmenti Anthony , Fleischer Arthur
Published: January 2005

Based upon data obtained with our arterio‐venous technique for the determination of cerebral metabolism in humans in vivo we have proposed a method for the determination of cerebral regional intermediary glucose metabolism in humans in vivo using specifically labeled 11C‐glucose and positron emission transverse tomography (PETT). In it we would give the subject successive intravenous injections of [3,4‐11C] glucose, [2,5‐11C] glucose and [1‐11C] glucose. There would be a 30 min period of continuous PETT measurements following each injection and a 2 hr interval after the first and second injections. The data would be used with suitable equations and algorithms to estimate for each specific region of the subject's brain the dynamics of the Embden‐Meyerhof‐Parnas (EMP) and the tricarboxylic acid cycle (TCA) metabolic pathways and the incorporation of glucose carbons into lactate, and the extent of dilution of glucose carbons in traversing the TCA with their subsequent incorporation into other carbon pools of the brain (ie, glutamate, glutamine, GABA, alanine). Using 14C as a model for 11C and autoradiographs made with rat brain slices, we have produced an animal model to demonstrate the feasibility of our proposed method. The resulting autoradiographs have provided evidence of the validity of the predictions made from our arterio‐venous data. The model was employed to show the selective reductions in the rates of incorporation of specific carbon atoms of glucose into regions of the rat brain and evidence of altered metabolic pathways following a single electroconvulsive shock (ECS) and after a series of nine ECS.
Decrease in the number of lower affinity (type II) nerve growth factor receptors on embryonic sensory neurons does not affect fiber outgrowth
Authors: Stach Robert W. , Wagner Barbara J.
Published: January 2005

Nerve growth factor binds to two different specific receptors on responsive cells. The relationship of these two receptors is not fully understood at this time. We have studied the binding of labeled NGF to a different strain of white leghorn chicken embryo dorsal root ganglionic cells. The equilibrium dissociation constants for the two sites (K = 4.1 ± 1.8 × 10−11M, K = 1.0 ± 0.8 × 10−9M) are identical to those obtained previously. Also, the number of type I sites per cell (3.8 ± 1.3 × 103) is the same as that previously determined. However, the number of type II sites per cell (1.9 ± 1.3 × 104) is significantly different than that previously determined. This 2.5‐fold decrease in the number of type II sites does not affect the concentration of NGF needed to obtain maximal fiber outgrowth from explanted sensory ganglia. The rate of association (1.2 ± 0.2 × 107 M−1 sec−1 at 22°C) of labeled NGF with receptors on sensory neurons from this different strain of chickens is identical to that previously obtained.

The rate of association of NGF with its receptors on sensory neurons was also determined at 4°C. This rate constant (2.1 ± 1.1 × 106 M−1 sec−1) along with the rate constants obtained at 22° and 37°C were used to determine an activation energy for the binding of NGF to its receptors. The activation energy obtained (16.2 kcal/mole) suggests that binding is not a diffusion‐controlled process.
Immunoselection of oligodendrocytes by magnetic beads. I. Determination of antibody coupling parameters and cell binding conditions
Authors: Meier D. H. , Lagenaur C. , Schachner M.
Published: January 2005

Oligodendrocytes from early postnatal mouse cerebellum were isolated using polyacrylamide‐coated magnetic beads carrying monoclonal antibody to 04 cell surface antigen. Oligodendrocytes were enriched to a purity of 91 ± 4% starting from a mixed cell population containing approximately 1.5% antigen‐positive oligodendrocytes. Viability of 04 antigen‐positive oligodendrocytes was approximately 90% as judged by exclusion of trypan blue. Oligodendrocytes were recovered after detachment from the beads with a yield of 19 ± 6% and after collection by centrifugation onto glass coverslips with yields of approximately 6% of all 04 antigen‐positive cells. The final cell yield of oligodendrocytes is approximately 8 × 105 cells/gram fresh cerebellar tissue.
Immunoselection of oligodendrocytes by magnetic beads. II. In vitro maintenance of immunoselected oligodendrocytes
Authors: Meier D. , Schachner M.
Published: January 2005

Oligodendrocytes isolated by immunoaffinity on magnetic beads using a monoclonal antibody to oligodendrocyte cell surfaces were obtained with a purity of 91 ± 4% [see companion paper, Meier et al, 1982]. Contaminating cells consisted mostly of tetanus toxin‐positive neurons with small cell bodies. Very few glial fibrillary acidic protein‐positive astrocytes and fibronectin‐positive fibroblasts or fibroblast‐like cells could be detected by indirect immunofluorescence. Macrophages, as identified by their characteristic morphology, were rarely detectable in the cultures. With increasing time in culture, the degree of homogeneity of the cultured oligodendrocytes increased slightly, while the cell density of oligodendrocytes decreased drastically within the first week of culture. Cultures of immunoselected oligodendrocytes with a lower degree of purity showed a better survival of oligodendrocytes, but a considerable reduction in purity after three weeks in culture, probably due to overgrowth of nonoligodendroglial elements.
Sequestration of tetanus toxin in developing neuronal cell cultures
Authors: Yavin Ziva , Yavin Ephraim , Kohn Leonard D.
Published: January 2005

Cultivation of pure populations of cerebral cells and the unambiguous identification of the neuronal and glial elements is a prerequisite for studying biochemical parameters associated with the developing nervous system under in vitro controlled conditions. We now report on sequestration of tetanus toxin, an exclusive marker for the neuronal cell surface, into a cellular compartment which prevents its release by neuraminidase. This compartment is associated with cellular elements remaining after detergent extraction of neuronal cells grown in monolayer cultures. Toxin sequestration into this compartment is preceded by a neuraminidase‐sensitive receptor mediated binding process which can be correlated with the appearance of lipid‐bound sialic acid in developing neuronal cultures.
Growth zone of the juvenile goldfish retina revealed by fluorescent flat mounts
Authors: Negishi K. , Teranishi T. , Kato S.
Published: January 2005

The retinas of juvenile goldfish (Carassius auratus) were processed for fluorescence microscope studies with flat mounts. The fish were reared in a laboratory aquarium for 1–3 months after either one class of retinal dopaminergic (DA) cells or indoleamine‐accumulating (IA) cells had been destroyed by intravitreal injection of 6‐hydroxydopamine (6‐OHDA; 2.5 m̈g) or 5,7‐dihydroxytryptamine (5,7‐DHT; 2.5 μg). The fish grew 10–12 cm in body length on the average for 3 months. The retinas were isolated at various survival times after pretreatment with 6‐OHDA or 5,7‐DHT. To improve detection of the cells, a mixture of noradrenaline (NA; 0.5 μg) and 5,6‐DHT (0.5 μg) was intravitreally injected into the eyes 2 hr before enucleation. Retinal growth occurred by cell proliferation at the circumferential margin distal to the retinal center. In this process, DA and IA cells were added in concentric rings, with a rate of cell accretion of approximately 12 cells per day, estimated from the mean cell density in the growth zone (about 252 μg width) over a 3‐month period. In order to compensate for loss of dopaminergic innervation induced by intravitreal 6‐OHDA, the proliferation rate appeared to be facilitated significantly compared to that in normal retinas, and long and centrally extending processes were found to emerge from the DA cells formed earliest after intravitreal injection of 6‐OHDA.
Regulation of neurite growth in purified retina neuronal cultures: Effects of PNPF, a substratum‐bound, neurite‐promoting factor
Authors: Adler Ruben
Published: January 2005

The responses of chick embryo retina neurons to the substratum‐bound, neurite‐promoting factor “PNPF” were studied using glia‐free, purified neuronal monolayers. Polyornithine‐coated dishes were exposed before cell seeding to either serum‐containing culture medium (PNPF(−) substratum) or to the same medium supplemented with 25% rat schwannoma conditioned medium, a source of PNPF (PNPF(+) substratum). The dishes were thoroughly rinsed before receiving a suspension of 8 day chick embryo neural retina cells in serum‐free medium.

The presence of PNPF on the substratum determined a dramatic increase in the relative frequency of neurite‐bearing cells in the cultures. After 6 hours in vitro PNPF(+) cultures contained 45% neurite‐bearing cells, as compared with 5–7% on PNPN(−) substrata. At 72 hours those values increased to 60% on PNPF(+) and to 40% on PNPF(−) substrata. PNPF(+) cultures also showed longer and/or more highly branched neurites, resulting in the formation of complex neurite networks. Moreover, a cell type characterized by the presence of a very long neurite could be seen on PNPF(+) but not on PNPF(−) substrata.

Six hour cultures were used to analyze in more detail the response of retinal neurons to PNPF. Addition of fetal calf serum to the medium determined a concentration‐dependent inhibition of neurite formation on PNPF(+) substrata. On the other hand, pretreatment of PNPF(+) substrata with concanavalin A also blocked the neurite‐promoting effect of the factor. This concentration‐dependent inhibitory effect of concanavalin A could be eliminated by the specific sugar α‐methyl‐D‐mannoside. Wheat germ agglutinin, another lectin known to react with PNPF, did not cause any reduction in the neurite‐promoting activity of this factor. Wheat germ agglutinin showed neurite‐promoting properties of its own in control experiments using PNPF(−) substrata.

The results indicate that the target spectrum of PNPF is broader than it was originally thought. Together with other reports from the literature, they also support the perception of neurite development as a cellular activity subject to complex regulatory mechanisms.
Two components of conditioned medium increase neuritic outgrowth from rat spinal cord explants
Authors: Dribin Lori B. , Barrett John N.
Published: January 2005

Conditioned medium (CM) from muscle or fibroblast cultures dramatically increases the outgrowth of neurites from fetal rat spinal cord slices in vitro. We report here that there are two separable fractions in conditioned medium that cause this increase in neuritic outgrowth. One fraction, with a molecular weight of approximately 50,000 daltons, enhances neuritic outgrowth only when it is present in the culture medium so that the slices are directly exposed to it. The second component has a much larger molecular weight (above 300,000 daltons), and can enhance neuritic outgrowth by directly binding to the culture substrate on pretreatment. Only when these two fractions are combined by pretreating the substrate with the higher molecular weight fraction and then growing the slices in the lower molecular weight fraction is the full outgrowth promoting activity of whole conditioned medium reconstituted. These two components act synergistically to reproduce nearly the full outgrowth promoting activity of non‐fractionated, whole conditioned medium.
Antiserum to a new neuronal growth factor: Effects on neurite outgrowth
Authors: Coughlin Michael D. , Kessler John A.
Published: January 2005

A new neuronal growth factor (CMF) isolated from mouse heart‐cell‐conditioned medium stimulates morphologic and biochemical development of mouse embryo sympathetic neurons [Coughlin et al, 1981]. Further analysis of CMF by chromatographic and electrophoretic procedures has shown that under nondissociating conditions, CMF activity is associated with very high molecular weight material. All biological activity eluted within the included volume of a Sepharose CL‐2B column in a molecular weight range corresponding to approximately 5 × 106 daltons. Similarly, electrophoresis showed no activity and very little protein entering 3% polyacrylamide gels, whereas both protein and activity migrated through 0.6% agarose‐1.2% acrylamide composite gels. To further characterize the biological effects of CMF on normal neuronal development, antibodies to CMF were employed. Rabbits immunized against CMF developed high titres of antibodies with activity specifically directed against CMF (anti‐CMF). Although anti‐CMF inhibited nerve growth factor (NGF)‐stimulated neurite outgrowth from the neonatal superior cervical ganglion, it did not inhibit the NGF‐stimulated increase in tyrosine hydroxylase activity. Moreover, ganglia incubated for 3 days in the presence of anti‐CMF were subsequently capable of producing neurites when washed and cultured in medium free of antiserum. Thus, anti‐CMF specifically blocked neurite extension without causing cell death or irreversible damage in ganglionic explants. Our observations suggest, therefore, that CMF or antigenically similar material is a requirement for neurite extension.
Developmental expression of rat brain mitogens for cultured astrocytes
Authors: Morrison Richard S. , Saneto Russell P. , de Vellis Jean
Published: January 2005

Extracts prepared from prenatal, neonatal, and adult rat brain were examined for the presence of astrocyte mitogens. Extracts were characterized by molecular weight separation on Sephadex G‐100. Two major peaks of mitogenic activity, at 80K and 23K daltons, were identified in the adult brain extract. Extract prepared from adult tissue also possessed the lowest specific activity. Extract prepared from neonatal rat brain exhibited four general peaks of activity at approximately 80K, 45K, 23K, and 17K daltons. This extract possessed the highest specific activity. Extract prepared from prenatal rat brain contained six general peaks of activity with unique peaks at 67K, 14K, and less than 10K daltons. In addition, this extract possessed a specific activity intermediate between the adult and postnatal extracts. Differences were also noted among the extracts in their requirement for “permissive” factors found in serum necessary for mitogenic activity. The data have implications for the normal development of astrocytes, as well as for the pathological process of gliosis.
Effect of epidermal growth factor on membrane motility and cell locomotion in cultures of human clonal glioma cells
Authors: Westermark Bengt , Magnusson Annika , Heldin Carl‐Henrik
Published: January 2005

Two clones, designated Cl 2 and Cl 3, were established from the human malignant glioma line U‐343 MGa. The astrocytic origin of the cells was proven by the presence in virtually 100% of the cells of the astrocyte marker glial fibrillary acidic protein. The addition of 10 ng epidermal growth factor (EGF) per milliliter to Cl 2 and Cl 3 cells resulted in the rapid appearance of large cell surface ruffles, visualized by scanning electron microscopy. A time course study by phase contract microscopy showed that the maximal ruffling activity occurred 5 minutes after addition of EGF. Under basic culture conditions (Eagle's MEM, 10% fetal calf serum), Cl 2 and Cl 3 cells were essentially immobile and formed tightly packed, wel demarcated colonies. In the presence of 10 ng EGF per milliliter, no defined colonies were formed and the cells seemed to move around freely. The stimulatory effect of EGF on cell migration was confirmed by growing the cells on a deposit of colloidal gold; in the absence of EGF, the cells remained immobile whereas cells grown at 10 ng EGF per ml formed long phagokinetic tracks. The effect of EGF on membrane motility and cell locomotion occurred in the absence of any effect of EGF on growth rate; both clones multiplied at the same rate in the absence as in the presence of EGF. Binding experiments using 125I‐labeled EGF demonstrated a single class of high affinity receptors. The number of 180,000 receptors per cell was estimated in both clones. The finding that human glioma cells in culture require EGF for their migration raises the interesting possibility that tumor cells in vivo may respond in a similar fashion, and in that case require a growth factor for migration and for the expression of their infiltrative growth potential. Furthermore, the present findings strengthen the notion that glial cells should be recognized as targets for EGF.
Effects of kainic acid injection and cortical lesion on ornithine and aspartate aminotransferases in rat striatum
Authors: Wong P. T.‐H. , McGeer E. G. , McGeer P. L.
Published: January 2005

The effects of cortical lesions and intrastriatal kainic acid injections on various striatal enzyme activities were investigated. Ornithine aminotransferase decreased concomitantly with glutamate uptake in decorticated and chronic kainic acid‐treated rats. It was also decreased in acute kainic acid‐lesioned striatum where glutamate uptake was unaffected. Aspartate aminotransferase, however, decreased only after acute kainic acid treatment. Results for glutamate uptake, glutamate decarboxylase, and choline acetyltransferase were in agreement with previous findings. These results suggest that ornithine may act as a precursor for glutamate in nerve terminals, although the nonspecific localization does not allow ornithine aminotransferase to be a convenient biochemical marker. The decrease in aspartate aminotransferase is thought to be due to be widespread cell degeneration after acute kainic acid. Aspartate aminotransferase activities were also found to be reduced in the frontal cortex, caudate nucleus and putamen of Huntington's disease brains.
Effects of the taurine transport antagonist, guanidinoethane sulfonate, and β‐alanine on the morphology of rat retina
Authors: Pasantes‐Morales H. , Quesada O. , Cárabez A. , Huxtable R. J.
Published: January 2005

Newborn rats treated for the first weeks of life with guanidinoethane sulfonate (GES), a blocker of taurine transport producing taurine depletion, showed a severe disruption of photoreceptor structure. Photoreceptor damage consisted of a marked reduction of the size of the photoreceptor layer, deformation of the outer segments, and a profound disorganization of the disc membranes. The GES‐induced degeneration pattern was very similar to that observed in cats fed a taurine‐deficient diet. Injection of β‐alanine, another antagonist of taurine transport, also produced a disruption of photoreceptor structure. These results confirm the requirement of taurine for maintaining photoreceptor structure in different species.