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At least 865 records · Page 48Linked to original sources
PubMed1993

Electron microscopic analysis of the rod pathway of the rat retina.

Two immunocytochemical markers were used to label the rod pathway of the rat retina. Rod bipolar cells were stained with antibodies against protein kinase C and AII-amacrine cells with antibodies against parvalbumin. The synaptic circuitry of rod bipolars in the inner plexiform layer (IPL) was studied. Rod bipolar cells make approximately 15 ribbon synapses (dyads) in the IPL. Both postsynaptic members of the dyads are amacrine cells; one is usually the process of an AII-amacrine cell and the other one frequently provides a reciprocal synapse. No direct output from rod bipolar cells into ganglion cells was found. AII-amacrine cells make chemical output synapses with cone bipolar cells and ganglion cells in sublamina a of the IPL. They make gap junctions with cone bipolar cells and other AII-amacrine cells in sublamina b of the IPL. The rod pathway of the rat retina is practically identical to that of the cat and of the rabbit retina. It is very likely that this circuitry is a general feature of mammalian retinal organization.

Animals↗
PubMed2004

In vivo analysis reveals different apoptotic pathways in pre- and postmigratory cerebellar granule cells of rabbit.

Naturally occurring neuronal death (NOND) has been described in the postnatal cerebellum of several species, mainly affecting the cerebellar granule cells (CGCs) by an apoptotic mechanism. However, little is known about the cellular pathway(s) of CGC apoptosis in vivo. By immunocytochemistry, in situ detection of fragmented DNA, electron microscopy, and Western blotting, we demonstrate here the existence of two different molecular mechanisms of apoptosis in the rabbit postnatal cerebellum. These two mechanisms affect CGCs at different stages of their maturation and migration. In the external granular layer, premigratory CGCs undergo apoptosis upon phosphorylation of checkpoint kinase 1 (Chk1), and hyperphosphorylation of retinoblastoma protein. In postmigratory CGCs within the internal granular layer, caspase 3 and to a lesser extent 7 and 9 are activated, eventually leading to poly-ADP-ribose polymerase-1 (PARP-1) cleavage and programmed cell death. We conclude that NOND of premigratory CGCs is linked to activation of DNA checkpoint and alteration of normal cell cycle, whereas in postmigratory CGCs apoptosis is, more classically, dependent upon caspase 3 activation.

Animals↗
PubMed2000

Crystallographic analysis of the reaction pathway of Zoogloea ramigera biosynthetic thiolase.

Biosynthetic thiolases catalyze the biological Claisen condensation of two acetyl-CoA molecules to form acetoacetyl-CoA. This is one of the fundamental categories of carbon skeletal assembly patterns in biological systems and is the first step in many biosynthetic pathways including those which generate cholesterol, steroid hormones and ketone body energy storage molecules. High resolution crystal structures of the tetrameric biosynthetic thiolase from Zoogloea ramigera were determined (i) in the absence of active site ligands, (ii) in the presence of CoA, and (iii) from protein crystals which were flash frozen after a short soak with acetyl-CoA, the enzyme's substrate in the biosynthetic reaction. In the latter structure, a reaction intermediate was trapped: the enzyme was found to be acetylated at Cys89 and a molecule of acetyl-CoA was bound in the active site pocket. A comparison of the three new structures and the two previously published thiolase structures reveals that small adjustments in the conformation of the acetylated Cys89 side-chain allow CoA and acetyl-CoA to adopt identical modes of binding. The proximity of the acetyl moiety of acetyl-CoA to the sulfur atom of Cys378 supports the hypothesis that Cys378 is important for proton exchange in both steps of the reaction. The thioester oxygen atom of the acetylated enzyme points into an oxyanion hole formed by the nitrogen atoms of Cys89 and Gly380, thus facilitating the condensation reaction. The interaction between the thioester oxygen atom of acetyl-CoA and His348 assists the condensation step of catalysis by stabilizing a negative charge on the thioester oxygen atom. Our structure of acetyl-CoA bound to thiolase also highlights the importance in catalysis of a hydrogen bonding network between Cys89 and Cys378, which includes the thioester oxygen atom of acetyl-CoA, and extends from the catalytic site through the enzyme to the opposite molecular surface. This hydrogen bonding network is different in yeast degradative thiolase, indicating that the catalytic properties of each enzyme may be modulated by differences in their hydrogen bonding networks.

Acetyl Coenzyme A↗
PubMed1999

Analysis of the folding pathway of chymotrypsin inhibitor by correlation of phi-values with inter-residue contacts

The transition state for folding of chymotrypsin inhibitor 2 (CI2) is investigated by correlating Phi-values with inter-residue contacts. In agreement with former work, the strongest consolidation of secondary structure is found in the alpha-helix. There are correlations for tertiary structure interactions between the residues Leu49, Ile57 and the helix which have been suggested to represent the main components of the nucleation site of CI2 folding. However, correlations for tertiary structure interactions of comparable magnitude are also found in the helix-strand2-strand1-motif and between strand3and strand4. Copyright 1999 Academic Press.

Journal Article↗
PubMed1990

Functional connections in the human temporal lobe. I. Analysis of limbic system pathways using neuronal responses evoked by electrical stimulation.

Connections in the human mesial temporal lobe were investigated using brief, single pulses of electrical stimulation to evoke field potential responses in limbic structures of 74 epileptic patients. Eight specific areas within these structures were stereotactically targeted for study, including amygdala, entorhinal cortex, presubiculum, the anterior, middle and posterior levels of hippocampus and the middle and posterior levels of parahippocampal gyrus. These sites were studied systematically in order to quantitatively assess the response characteristics and reliability of responses evoked during stimulation of pathways connecting the areas. Specific measures included response probability, amplitude, latency and conduction velocities. The results are assumed to be representative of typical human limbic pathways since all recordings were made interictally and response probabilities across sites were not found to differ significantly between non-epileptogenic vs. identified epileptogenic regions. Field potentials ranging in amplitude from less than 0.1 to greater than 6.0 mV were evoked ipsilaterally, with mean onset latencies and conduction velocities ranging from 4.4 ms and 3.64 m/s in the perforant pathway connecting entorhinal cortex to anterior hippocampus to 24.8 ms and 0.88 m/s in the pathway connecting the amygdala and middle hippocampus. Stimulation of presubiculum and entorhinal cortex were most effective in evoking widespread responses in adjacent limbic recording sites, whereas posterior parahippocampal gyrus appeared functionally separated from other limbic sites since its probability of influencing ipsilateral sites was significantly lower than any other area. It was particularly noteworthy that stimulation did not evoke responses in any sites in contralateral hippocampal formation; even though a large number of sites were tested with bilateral implantation of homotopic electrodes.(ABSTRACT TRUNCATED AT 250 WORDS)

Action Potentials↗
PubMed1993

Pentose-phosphate pathway in Saccharomyces cerevisiae: analysis of deletion mutants for transketolase, transaldolase, and glucose 6-phosphate dehydrogenase.

Deletion mutants for the yeast transketolase gene TKL1 were constructed by gene replacement. Transketolase activity was below the level of detection in mutant crude extracts. Transketolase protein could be detected as a single protein band of the expected size by Western-blot analysis in wild-type strains but not in the deletion mutant. Deletion of TKL1 led to a reduced but distinct growth in synthetic medium without an aromatic amino-acid supplement. We also isolated double and triple mutants for transketolase (tkl1), transaldolase (tal1), and glucose 6-phosphate dehydrogenase (zwf1) by crossing the different mutants. A tal1 tkl1 double mutant grew nearly like wild-type in rich medium. Only the tkl1 zwf1 double and the tal1 tkl1 zwf1 triple mutant grew more slowly than the wild-type in rich medium. This growth defect could be partly alleviated by the addition of xylulose but not ribose. The triple mutant still grew slowly on a synthetic mineral salts medium without a supplement of aromatic amino acids. This suggests the existence of an alternative but limited source of pentose phosphates and erythrose 4-phosphate in the tkl1 zwf1 double mutants. Hybridization with low stringency showed the existence of a sequence with homology to transketolase, possibly a second gene.

Base Sequence↗
PubMed1980

Analysis of the fermentation pathways of clostridia using double labelled glutamade.

L-(4-14C, 3-3H)Glutamate was used as a tool to elucidate the pathway of its fermentation. The methylaspartate pathway was found in strains of Clostridium tetanomorphum, C. malenominatum, C. limosum, C. lentoputrescens and C. tetani, whereas the hydroxyglutarate pathway was detected in C. sporosphaeroides. These data were supported by measurement of enzyme activities present in cell-free extracts.

Aspartic Acid↗
PubMed1976

The peristaltic reflex: an analysis of the nerve pathways and their pharmacology.

The enteric reflexes in isolated segments of the distal colon and rectum of the guinea-pig were studied by applying localized distensions and recording the consequent changes in circular muscle activity, and by recording tension changes in the circular muscle during the propulsion of a bolus in vitro. Lesions of the wall of the colon were made to locate nerve pathways involved in the reflexes and pharmacological tests were applied to investigate the natures of transmitters released and the types of receptors involved. Distension produced a transient contraction of the circular muscle on the oral side and sustained relaxation on the anal side. Both reflexes were nerve-mediated. They were elicited in segments deprived of mucosa and submucosa. Interruption of Auerbach's plexus, but not interruption of the submucosal plexus, prevented their conduction. The ascending excitatory reflex was partly blocked by hyoscine and was also partly blocked by methysergide or by making the preparation tachyphylactic to the excitatory action of 5-hydroxytryptamine. The ascending excitatory pathways apparently involve neurons releasing a 5-HT-like transmitter as well as cholinergic neurons. The descending inhibitory reflex was not antagonized by hyoscine, guanethidine, methysergide or mepyramine. It is assumed that the inhibitory neurons activated in this reflex are identical with the non-cholinergic, non-adrenergic, enteric inhibitory neurons found throughout the intestine. If both the ascending excitatory and descending inhibitory reflexes acted simultaneously on the same area of circular muscle, the inhibitory response tended to dominate. Pellets of faeces, covered by a thin layer of resin, were introduced into the oral ends of isolated segments of colon. They were propelled analwards at speeds of 0.5-1.6 mm/s. Tension records showed that the pellets were preceded by relaxation and followed by a ring of contraction. The propulsion was blocked by both hyoscine and methysergide. Descending waves of contraction were also observed in empty segments of colon. These occurred spontaneously or were initiated by stretch. They did not occur in the presence of hyoscine or tetrodotoxin. It is postulated that three factors may contribute to propulsion in the guinea-pig distal colon: ascending excitatory reflexes which evoke contractions above a bolus; descending inhibitory reflexes which cause relaxations below; and contractions which, once set up in the circular muscle, travel in an anal direction.

Animals↗
PubMed1987

A cell-free analysis of the endocytic pathway.

The molecular control of the endocytic pathway is poorly understood. To obtain this information requires the use of cell-free systems which faithfully recreate the various endocytic events as they occur in the intact cell. Here I describe our approach to elucidating the mechanism which controls the fusion between different vesicles on the pathway.

Animals↗
PubMed2005

Analysis of signal transduction pathways in macrophages using expression vectors with CMV promoters: a cautionary tale.

The cytomegalovirus (CMV) major immediate-early promoter is a strong promoter used for both in vitro and in vivo expression of proteins in signal transduction and gene therapy studies. CMV activity is induced by external stimuli such as endotoxin from Gram-negative bacteria (LPS), TNF-alpha and phorbol esters. This inducibility poses problems when this promoter is used to drive the expression of either wild type or dominant negative mutated proteins as tools in signal transduction studies. This report draws attention to the problem associated with this widely used approach. The role of NF-kappaB and Hypoxia Inducible Factor-1alpha (HIF-1alpha) in the transcriptional regulation of Vascular Endothelial Growth Factor (VEGF) in macrophages was investigated using CMV-promoter-driven expression of either wild type or dominant negative proteins involved in these pathways. Difficulties encountered while interpreting the data due to the inducibility of the CMV promoter by LPS are highlighted in this report and provide a cautionary note for the evaluation of data acquired using this approach.

Animals↗
PubMed1992

Suppression of nociceptive responses in parafascicular neurons by stimulation of substantia nigra: an analysis of related inhibitory pathways.

A total of 166 neurons in parafascicular nucleus (PF) were studied, 85 from intact animals, 72 following dorsal spinal cord transection (D.Sp.C.X.), and 9 following complete transection of the spinal cord. Two patterns of nociceptive responses were identified following noxious stimulation and these responses were classified as 'nociceptive-on' and 'nociceptive-off' neurons, respectively. The effects of stimulating the substantia nigra (SNS) on the spontaneous and on the nociceptive evoked discharges were observed and compared in intact, D.Sp.C.X. and completely transected spinal cord rats. The results show that SNS significantly suppresses both the spontaneous and the nociceptive evoked discharges elicited by peroneal nerve stimulation. With an intact spinal cord, SNS suppressed both the spontaneous [-37 +/- 3.2% (P less than 0.05)] and the nociceptive evoked discharges [-52.8 +/- 2.8% (P less than 0.01)] of the 'nociceptive-on' cells respectively, while in the 'nociceptive-off' cells the same stimulation elicited an even more prominent suppression upon both discharges (-47.7 +/- 5.4%, P less than 0.01 and -64.9 +/- 5.0%, P less than 0.01), respectively. After D.Sp.C.X., the suppressive effects on the 'nociceptive-on' cells following SNS were diminished (-28.1 +/- 3.5% and -36.9 +/- 2.6%, respectively) but not abolished, while in the 'nociceptive-off' cells, the inhibitory effects on SNS were unchanged. In addition, the suppressive effects of SNS on the spontaneous activity of PF neurons in cases with completely cut spinal cords remains unchanged. These results suggest that SNS modulates the spontaneous and the noxious evoked responses of the PF neurons by way of supraspinal connections besides the previously described descending projecting pathways.

Action Potentials↗