Scanning Electron Microscopy of Xiphinema,Longidorus, and Californidorus Stylet Morphology

Stylet ultrastructure of five Xiphinema, four Longidorus, and three Californidorus species was compared by scanning electron microscopy. Morphological differences were seen in the odontophores and odontostyle bases between the genera and some of the species. All Xiphinema studied had well-developed odontophore flanges; the Longidorus species lacked flanges, except for weakly developed ones in L. diadecturus; and none of the Californidorus had flanges. Three sinuses were present in the odontophores of all species. The sinuses varied in length depending upon species. In Xiphinema and Californidorus the odontostyle bases had distinct overlapping collars, but in Longidorus the collars were absent except for L. diadecturus. The odontostyle-odontophore junction from a lateral view appeared as a slanted transverse line in all the species, but in a dorsal view of Xiphinema and Californidorus it was V-shaped. Dorsal longitudinal seams of the odontostyle and odontophore were observed in all the species. The dorsally located odontostyle aperture was ca. 1 μm from the anterior end in all species, except in one Longidorus sp. it was ca. 4 μm from the end.     

Analysis of Adenosine Immunoreactivity, Uptake, and Release in Purified Cultures of Developing Chick Embryo Retinal Neurons and Photoreceptors

We have investigated the presence of endogenous adenosine and of mechanisms for adenosine uptake and release in chick embryo retinal neurons and photoreceptors grown in purified cultures in the absence of glial cells. Simultaneous autoradiographic and immunocytochemical analysis showed that endogenous adenosine and the uptake mechanism for this nucleoside colocalize in practically all the photoreceptors, but only in approximately 20% of the neurons. Approximately 25% of the neurons showed either immunocytochemical labeling or autoradiographic labeling, while greater than 50% of the neurons were unlabeled with both techniques. [3H]Adenosine uptake was saturable and could be inhibited by nitrobenzylthioinosine and dipyridamole and by pretreatment of the [3H]adenosine with adenosine deaminase. Although these observations indicate that the uptake is specific for adenosine, only 35% of accumulated radioactivity was associated with adenosine, with the remaining 65% representing inosine, hypoxanthine, and nucleotides plus uric acid. Adenosine as well as several of its metabolites were released by the cells under basal as well as K(+)-stimulated conditions. Potassium-enhanced release was blocked by 10 mM CoCl2 or in Ca2(+)-free, Mg2(+)-rich solutions. The results indicate that retinal cells that synthesize, store, and release adenosine differentiate early during embryogenesis and are therefore consistent with a hypothetical role for adenosine in retinal development.     

Development of Bursaphelenchus xylophilus Populations in Wood Chips with Different Moisture Contents

Bags of Pinus strobus wood chips with moisture contents of 38, 92, 164, and 217% (oven dry weight) were inoculated with Bursaphelenchus xylophilus and incubated at 30 C in order to determine the effect of wood moisture on nematode population development. Nematodes were extracted after 2, 4, 8, and 12 weeks. Population levels were greatest in wood chips with a moisture content of 38% and decreased successively with each higher moisture content. In chips with the three lower moisture contents, populations peaked at 2 weeks, but at 217% moisture, they peaked at 8 weeks. By 12 weeks, nematode populations had declined in wood chips with 92 and 164% moisture contents. The fungi most frequently isolated from the wood chips were Alternaria, Fusarium, Gliocladium, Graphium, Penicillium, Trichoderma, and Mucorales.     

Decreased Protein Kinase C Activity During Cerebral Ischemia and After Reperfusion in the Adult Rat

The possible activation of protein kinase C (PKC) during total cerebral ischemia was investigated in the rat. Translocation of PKC activity from the soluble to the particulate fraction was used as an index of PKC activation. There was a drop in the proportion of particulate PKC activity from 30% for controls to 20% by 30 min of ischemia (p less than 0.01). By 20 min of cardiac arrest, there was a 40% decline of the total cellular PKC activity (p less than 0.01). This was not accompanied by an increase in activator-independent activity, a finding indicating PKC was not being converted to protein kinase M. These data suggest that PKC was not activated during ischemia, but rather that ischemia causes a reduction in cellular PKC activity. Translocation of PKC activity to the particulate fraction was not observed in the cerebral cortex or hippocampus of reperfused brain for up to 6 h of recovery following 11-13 min of total cerebral ischemia. The level of total, soluble, and particulate PKC activity in the cerebral cortex was reduced (p less than 0.05), corresponding to the decrease observed by 15 min of ischemia without reflow. A similar decline in activity was also observed in the hippocampus. No increase in activator-independent activity was observed. These data suggest that PKC was inhibited during cerebral ischemia and that this reduced level of PKC activity was maintained throughout 6 h of recovery. We conclude that pathological activation of PKC was not responsible for the evolution of ischemic brain damage.     

Modulation of Opioid Receptor Binding by Cis and Trans Fatty Acids

In synaptosomal brain membranes, the addition of oleic acid (cis), elaidic acid (trans), and the cis and trans isomers of vaccenic acid, at a concentration of 0.87 mumol of lipid/mg of protein, strongly reduced the Bmax and, to a lesser degree, the binding affinity of the mu-selective opioid [3H]Tyr-D-Ala-Gly-(Me)Phe-Gly-ol ([3H]DAMGO). At comparable membrane content, the cis isomers of the fatty acids were more potent than their trans counterparts in inhibiting ligand binding and in decreasing membrane microviscosity, both at the membrane surface and in the core. However, trans-vacenic acid affected opioid receptor binding in spite of just marginally altering membrane microviscosity. If the receptors were uncoupled from guanine nucleotide regulatory protein, an altered inhibition profile was obtained: the impairment of KD by the fatty acids was enhanced and that of Bmax reduced. Receptor interaction of the delta-opioid [3H](D-Pen2,D-Pen5)enkephalin was modulated by lipids to a greater extent than that of [3H]DAMGO: saturable binding was abolished by both oleic and elaidic acids. The binding of [3H]naltrexone was less susceptible to inhibition by the fatty acids, particularly in the presence of sodium. In the absence of this cation, however, cis-vaccenic acid abolished the low-affinity binding component of [3H]naltrexone. These findings support the membrane model of opioid receptor sequestration depicting different ionic environments for the mu- and delta-binding sites. The results of this work show distinct modulation of different types and molecular states of opioid receptor by fatty acids through mechanisms involving membrane fluidity and specific interactions with membrane constituents.     

Effect of Desipramine on a Glycoprotein Sialyltransferase Activity in C6 Cultured Glioma Cells

The tricyclic antidepressant desipramine, when added to culture medium, gave rise in C6 rat glioma cells to a decrease of the activity of the enzyme asialofetuin sialyltransferase. The inhibition was dose and time dependent and was observed in both multiplying cells and cells blocked with 2 mM thymidine or depletion of amino acids. This inhibition was rather specific to the sialyltransferase, as under the conditions where this enzyme was inhibited up to 70%, other enzymes such as dolichol phosphate mannose synthetase, glutamine synthetase, and glycerol phosphate dehydrogenase remained unaffected. This inhibition was not reversed after removal of desipramine from the medium and was not observed by direct addition of desipramine to the sialyltransferase incubation assay. Under the same conditions, W-7 [N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide], which is known to be a potent calmodulin antagonist and an inhibitor of calmodulin-dependent kinases, gave the same concentration-dependent inhibition profile of sialyltransferase as desipramine, whereas H-7 [1-(5-isoquinolinylsulfonyl)-2-methylpiperazine], which is an inhibitor of protein kinase C and cyclic nucleotide-dependent kinases, had no effect. So, it is suggested that desipramine inhibits the sialyltransferase activity in C6 glioma cells through a calmodulin-dependent system.     

Location in Muscarinic Acetylcholine Receptors of Sites for [3H]Propylbenzilcholine Mustard Binding and for Phosphorylation with Protein Kinase C

Muscarinic acetylcholine receptors purified from porcine cerebra or atria were covalently labeled with [3H]propylbenzilylcholine mustard ([3H]PrBCM), and then the labeled receptors were subjected to limited hydrolysis with trypsin, V8 protease, and lysyl endopeptidase, followed by analysis involving sodium dodecyl sulfate-polyacrylamide gel electrophoresis, fluorography, autoradiography, or immunostaining. The labeled peptides were located on the basis of their reactivity with antibodies raised against three synthetic peptides with partial sequences of the m1 or m2 receptor, and of their sensitivity to endoglycosidase F, which was taken as evidence that they contain glycosylation sites near the N terminus. The [3H]PrBCM-binding site in both cerebral and atrial receptors was found to be located between the N terminus and the second intracellular loop, because the size of the smallest deglycosylated peptide that contained both the [3H]PrBCM-binding and glycosylation sites was approximately 16 kDa. Cerebral receptors were 32P-phosphorylated with protein kinase C, and the major phosphorylation sites in cerebral muscarinic receptors were found to be located in a C-terminal segment including a part of the third intracellular loop, because a 32P-labeled peptide of 12-14 kDa reacted with anti-(m1 C-terminal peptide) antiserum. The presence of an intramolecular disulfide bond, probably between Cys 98 and Cys 178 in the first and second extracellular loops, respectively, was suggested by the finding that a peptide of approximately 17 kDa containing the [3H]PrBCM-binding site, but not the glycosylation sites, was partly converted to a peptide of approximately 12 kDa on treatment with beta-mercaptoethanol.     

Labeling of Retina and Optic Tectum Phospholipids in Chickens Exposed to Light or Dark

The labeling of retina ganglion cell and optic tectum phospholipids was determined in chickens given an intraocular injection of 32P and then either exposed to light or maintained in the dark. Significantly higher labeling was found in the optic tectum phospholipids of light-exposed compared with dark-maintained animals after 3-24 h of labeling. In the ganglion cells, the labeling of phospholipids increased in dark with respect to light at 15 and 30 min of labeling; from 60 min to 24 h, the labeling of phospholipids was significantly higher in light with respect to dark, even if the precursor pool showed a higher labeling in dark at all times studied. When labeling was allowed to proceed in the dark for 30 min and then half of the animals were exposed to light for 15 min, the labeling of ganglion cell phospholipids of light-exposed animals was significantly higher than those of animals kept in the dark. No individual phospholipid accounted for the differences observed in the labeling of the total phospholipid pool. These results are interpreted as an increase in the biosynthesis of phospholipids in the ganglion cell somas in light with respect to dark.     

Transport of Tryptophan into Brain from the Circulating, Albumin-Bound Pool in Rats and in Rabbits

Tryptophan is the only amino acid in the circulation that is bound by albumin, and previous studies have suggested that the brain tryptophan supply is a function of either the free or the albumin-bound pool of tryptophan in blood. Since the albumin molecule per se does not cross the brain capillary wall, i.e., the blood-brain barrier (BBB), the transport of tryptophan from the circulating albumin-bound pool may involve enhanced dissociation of tryptophan from the albumin binding sites within the cerebral microcirculation. This hypothesis was confirmed in the present studies wherein the dissociation constant (KaD) of albumin binding of tryptophan in the rat or rabbit brain microcirculation was measured in vivo. Brain extraction data for [14C]tryptophan determined with the carotid artery injection technique were fit to the Kety-Renkin-Crone equation modified for protein-bound solute. The KaD of albumin binding in the rat or rabbit brain microcirculation under pentobarbital anesthesia was 1.7 +/- 0.1 and 3.9 +/- 1.0 mM, respectively, as compared to the KD value measured in vitro with equilibrium dialysis, 0.13 +/- 0.03 mM. In contrast, the KaD value of albumin binding of tryptophan in vivo in the rabbit brain microcirculation was reduced by ether anesthesia to a value of 2.1 +/- 0.4 mM. This reduction in the KaD under ether anesthesia was associated with a 2.5-fold increase in cerebral blood flow. In addition, dialyzed rabbit serum caused a statistically significant inhibition in [14C]tryptophan influx during ether, but not pentobarbital, anesthesia.(ABSTRACT TRUNCATED AT 250 WORDS)