Biomedical subjects

R E Anderson

Publications and source records attributed to R E Anderson.

At least 19 recordsLinked to original sources

PubMed1992

Activation of bovine rod outer segment phospholipase C by arrestin.

Phospholipase C (PLC) enzyme activity in rod outer segment (ROS) membranes bleached in the presence of ATP and GTP was assayed using exogenously added [3H]phosphatidylinositol 4,5-bisphosphate vesicles as substrate. The addition of the soluble ROS protein arrestin (also known as S-antigen or 48K protein) to ROS membranes activated PLC 2-3.4-fold. This activation was dose-dependent, and maximal activation was observed at an arrestin concentration of congruent to 110-220 nM. PLC activation by arrestin was dependent on ROS protein concentration and free Ca2+. Soluble PLC (s-PLC) enzyme activity present in hypotonic extracts of bleached ROS was also activated 2-4-fold by arrestin. Maximum activation of s-PLC by arrestin was observed at free Ca2+ of 80 nM. Arrestin activation of s-PLC was not affected by urea-treated and extensively washed ROS membranes, suggesting that rhodopsin was not required for the observed effect of arrestin on s-PLC. The results are indicative of a direct interaction of arrestin with s-PLC, resulting in the activation of the latter. Based on these results and the documented binding of arrestin to bleached and phosphorylated rhodopsin, a model for the light activation of PLC in ROS is proposed.

Animals↗

PubMed1992

Quantitation of phenacyl esters of retinal fatty acids by high-performance liquid chromatography.

A high-performance liquid chromatographic (HPLC) method for the separation and quantitation of retinal fatty acids containing long-chain polyunsaturated fatty acids is described. Fatty acids from frog retinal lipids were converted to the corresponding phenacyl derivatives which were separated on a C18 reversed-phase column and detected at 242 nm. Molar absorptivities (peak area units/nmol) of up to seventeen fatty acid phenacyl derivatives were determined and used for quantitation of fatty acids separated by HPLC. Compared with gas chromatography, the HPLC method gave a similar molar percent distribution of the fatty acids and was twenty to fifty times more sensitive. This HPLC method provides a useful means for the study of chemistry and metabolism of long-chain polyunsaturated fatty acids in retina and other tissues where amounts of material may be limited or recovery of individual components desirable.

Acetophenones↗

PubMed1992

Conservation of docosahexaenoic acid in the retina.

Over the last several years, evidence has accumulated that n-3 fatty acids, particularly 22:6n-3, are essential for the development of the structure and function of the visual system. The importance of 22:6n-3 is reflected in the tenacious manner in which the retina conserves this fatty acid during n-3 deficiency. We have shown that conservation is achieved by recycling 22:6n-3 within the retina or between the retina and the pigment epithelium. Within the retina, recycling could be accomplished by deacylation-reacylation reactions (Louie et al., 1991; Zimmerman and Keys, 1988). Recycling between the retina and the RPE may be achieved through specific transport proteins, possibly interphotoreceptor retinoid-binding protein (Bazan et al., 1985) and/or apolipoprotein E (Bazan et al., 1991).

Animals↗

PubMed1992

Docosahexaenoic acid increases in frog retinal pigment epithelium following rod photoreceptor shedding.

The vertebrate retina conserves docosahexaenoic acid (22:6n-3) during n-3 fatty acid deficiency. The mechanism of conservation is not known, although recycling of this fatty acid between the retinal pigment epithelium (RPE) and retina is one possibility. We examined the role of the RPE in conservation of 22:6n-3 by quantitating the fatty acids and phospholipid molecular species (PLMS) in frog RPE before and after light-stimulated shedding of rod outer segments (ROS). RPE cells were dissociated with brush agitation and purified by a discontinuous ficoll density gradient. One hour after the light-induced shedding of ROS, the phagocytosed ROS tip and opsin content of RPE had increased. Simultaneously, the levels of 22:6n-3 and 22:6(n-3)-containing PLMS were increased in the RPE. Within 8 hr following the shedding event, 22:6n-3 in the RPE had returned to the dark level. These findings indicate that the phagocytosed ROS tips contain 22:6n-3 and that the RPE metabolizes these ROS tips and eliminates 22: 6n-3 from the cell. Thus, the RPE is intimately involved in the metabolism of 22: 6n-3 in the retina. The recycling of 22: 6n-3 from the RPE to the retina is a possible means of conserving this important fatty acid in the retina.

Animals↗

PubMed1992

Decreased docosahexaenoic acid levels in retina and pigment epithelium of frogs fed crickets.

Whole retina, rod outer segments, and retinal pigment epithelium of frogs (Rana pipiens) fed crickets for more than 1 year had significantly lower levels of docosahexaenoic acid (22: 6n-3) than the same tissues of frogs fed crickets for less than 1 month. Decreases in 22:6n-3 levels in these tissues were compensated for by increases in the n-6 polyunsaturated fatty acids (PUFAs), primarily 22:5n-6. There were no changes in the levels of saturated, monoenoic, or dienoic acids. Analysis of diacyl phospholipid molecular species (PLMS) revealed decreases in both the 22:6(n-3)-containing dipolyenoic molecular species in phosphatidylethanolamine and phosphatidylserine, and the monopolyenoic molecular species in phosphatidylcholine. These PLMS were replaced by species containing 22:5n-6 or other n-6 PUFAs. Examination of fatty acid methyl esters of total lipids extracted from crickets revealed that less than 1 mol% fatty acids were of the n-3 family, while more than 30 mol% were of the n-6 family. Thus, frogs raised on an n-3-deficient diet have reduced levels of n-3 PUFA in their retinas, rod outer segments, and retinal pigment epithelium. Although such changes have been reported for mammals, this is the first report of the effects of n-3 deficiency on the lipids of amphibians.

Animals↗

PubMed1992

Uptake of 22-carbon fatty acids into rat retina and brain.

Rat retina accumulates high levels of 22-carbon (C22) polyunsaturated fatty acids (PUFA), especially docosahexaenoic acid (DHA, 22:6 omega 3), in rod outer segment (ROS) phospholipids (PL). However, plasma, the source of retina lipids, is enriched in 20-carbon (C20) fatty acids instead of C22 PUFA. This suggests that the retina has a mechanism(s) for selective uptake of C22 PUFA from the blood. It is not known if the selective uptake is specific for the carbon number alone, or if the number of double bonds is also important. To address this question, the following study was carried out using erucic acid (22:1 omega 9) as a metabolic marker molecule. Albino rats were raised from birth on a diet containing 10% (by weight) of either rapeseed oil (43% 22:1 omega 9) or blended canola oil (0.4% 22:1 omega 9). At 4 months of age, plasma, liver, adrenal gland, brain and retina were collected, lipids were extracted, and fatty acids were determined. In those rats fed rapeseed oil, 22:1 omega 9 was incorporated into the lipids of plasma (2.3%), liver (0.6%), and adrenal gland (17.6%), indicating that this fatty acid was absorbed, transported, and metabolized by the rats. However, 22:1 omega 9 was not incorporated into the lipids of retinal ROS or brain. Our results suggest that both the carbon number and degree of unsaturation are important determinants in the selective uptake of C22 fatty acids from plasma into both the brain and the retina.

Animals↗

PubMed1992

Panoramic imaging of brain pHi and CBF during penicillin and metrazole induced status epilepticus.

Using real-time in vivo umbelliferone fluorescent imaging, cortical intracellular brain pH (pHi) and cortical blood flow (CBF) were measured in New Zealand white rabbits during generalized seizures induced by intravenous metrazole or sodium penicillin. In the former, brain pHi declined from 7.04 +/- 0.07 to 6.78 +/- 0.07 within 15 min of generalized seizures and remained at this level for 1 h. In the penicillin group, pHi fell from 7.05 +/- 0.10 to 6.81 +/- 0.07 and also remained at this level over 60 min. This brain acidosis was uniform across the brain's surface. With the onset of status epilepticus there was a hyperemia which occurred in a heterogeneous pattern with blood flow appearing to be greater adjacent to cortical vasculature and slower in border zones between surface blood vessels. In the metrazole group, there was evidence of vasomotor paralysis with loss of autoregulation involving both cortical surface vasculature and penetrating arterioles with their capillary beds.

Animals↗

PubMed1992

Focal cortical distribution of blood flow and brain pHi determined by in vivo fluorescent imaging.

A technique was developed for panoramic imaging of intracellular focal brain pH (pHi) and focal cortical blood flow to measure the pattern of changes in these parameters during arterial PCO2 (PaCO2) and arterial PO2 (PaO2) alterations. Eleven overnight-fasted New Zealand White rabbits were operated and studied under 2.0 and 1.0% halothane anesthesia, respectively. The exposed cortex was imaged by a 512 x 512 pixel image intensifier system with a resolution of 10 microns2/pixel. Focal brain pHi and focal cortical blood flow were measured by umbelliferone fluorescence. The mean focal cortical blood flow was 49 +/- 4.5 ml.100 g-1.min-1, and focal brain pHi was 7.05 +/- 0.02 during normocapnia. At a stable PaCO2, focal cortical blood flow and focal brain pHi within 10 microns of surface-conducting vessels were 2-50% greater and 0.02 pH units lower than that observed in the border-zone regions between vessels. Focal cortical blood flow demonstrated a significant heterogeneity of 51% among various cortical regions, despite a stable PaCO2. Alternatively, focal brain pHi was homogeneous, varying no more than 0.05 pH units. These results demonstrate the dependence of focal cortical blood flow on topical locations from surface conducting vessels and the tight regulation of brain pHi.

Animals↗

PubMed1992

Enrichment of polyunsaturated fatty acids from rat retinal pigment epithelium to rod outer segments.

Polyunsaturated fatty acids (PUFA), especially docosahexaenoic acid (DHA, 22:6n-3), are enriched in phospholipids of vertebrate rod outer segments (ROS). Retinal ROS can incorporate 22 carbon (C-22) PUFA from the plasma pool where C-20 PUFA are predominant. In this study, we analyzed the fatty acid composition of retinal pigment epithelium (RPE) and ROS from rats fed different fatty acid supplements to determine whether this enrichment is at the photoreceptor-RPE boundary or the RPE-choriocapillaris boundary. Long Evans rats were raised from birth for 13-14 weeks on a diet supplemented with 10% (wt/wt) hydrogenated coconut oil (COC; 0.2% 18:2n-6, no 18:3n-3), safflower oil (SAF; 73.8% 18:2n-6, 0.1% 18:3n-3), or linseed oil (LIN; 16.4% 18:2n-6, 52.2% 18:3n-3). These diets were chosen because they increased plasma levels of 20:3n-9, 20:4n-6, and 20:5n-3, respectively. These three fatty acids served as metabolic markers. Plasma levels of 22:6n-3 were reduced by the COC and SAF diets. The RPE incorporated 20:3n-9, 20:4n-6, and 20:5n-3 from the plasma. However, the levels of 20:3n-9 and 20:5n-3 were very low in ROS and 20:4n-6 was not significantly elevated in the ROS of the SAF diet group. The relative amount of total C-20 PUFA in phospholipids in RPE was similar to that found in plasma and was about 4-16 times (depending on different lipid classes) that in the ROS. In contrast, C-22 PUFA (22:6n-3 and 22:5n-6) showed a step-wise, average 3-5 fold increase in concentration from the plasma to the RPE to the ROS.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Lipids of frog retinal pigment epithelium: comparison with rod outer segments, retina, plasma and red blood cells.

The glycerolipid and fatty acid compositions of frog retinal pigment epithelium (RPE) were determined and compared with rod outer segments (ROS), retina, plasma, and red blood cells (RBC). The glycerolipid class composition of RPE was similar to RBC and ROS or retina, with phosphatidylcholine and phosphatidylethanolamine being the major components. The fatty acid composition of RPE differed substantially from that of plasma or RBC; the former contained much higher levels of C-20 and C-22 polyunsaturated fatty acids (PUFAs), such as 20:4n-6 and 22:6n-3, but less C-18 mono-, dienoic, and trienoic acids. The difference between RPE and ROS or retina with respect to fatty acid profile was also dramatic; RPE had relatively less 22:6n-3, but more 20:4n-6 and 18:2n-6, than ROS or retina. These results suggest that frog RPE cells may selectively take up C-20 and C-22 PUFAs from the circulation, but preferentially deliver 22:6n-3 to the ROS and retina. Fatty acid analyses show that 20:4n-6 and 22:6n-3 were unevenly distributed among RPE glycerolipids; phosphatidic acid, diglyceride, triglyceride, and phosphatidylserine are relatively more enriched in 22:6n-3 compared with 20:4n-6. This information might imply that these two PUFAs are metabolized differently inside the frog RPE cells.

Animals↗

PubMed1992

Effects of intermittent reperfusion during temporal focal ischemia.

There is controversy regarding the role of intermittent reperfusion employed as a cerebroprotective measure when temporary arterial occlusion is necessary during repair of difficult aneurysms. The intraluminal suture middle cerebral artery (MCA) occlusion technique was used in 23 Wistar rats under barbiturate anesthesia to induce 60, 90, or 120 minutes of uninterrupted MCA occlusion. The total infarcted areas obtained were compared to those occurring in 27 animals subjected to identical cumulative ischemic periods but with 5 minutes of reperfusion after every 10-minute ischemic period. The mean total infarcted areas in the groups with 60-minute (1.8 +/- 0.89 sq mm), 90-minute (1.08 +/- 1.02 sq mm), and 120-minute (8.72 +/- 5.89 sq mm) intermittent reperfusion were significantly smaller than those occurring in the 60-minute (12.02 +/- 3.10 sq mm), 90-minute (11.54 +/- 2.68 sq mm), or 120-minute (30.43 +/- 6.51 sq mm) control groups, respectively (p < 0.05). Furthermore, there was no difference in the occurrence of blood-brain barrier breakdown, intraparenchymal hemorrhage, hemispheric edema, or seizures between control and intermittent reperfusion groups. The results support the hypothesis that intermittent reperfusion is beneficial if vessel occlusion is required during aneurysm repair.

Animals↗

PubMed1992

Region and age-dependent variation in susceptibility of the human retina to lipid peroxidation.

Trephined buttons from the posterior central (including the macula) and four peripheral regions of human retina were subjected to in vitro lipid peroxidation initiated by Fe+2. There was an age-related increase in peroxidation in the retinal tissue from the posterior region (P = 0.0019), but not in tissues from any of the four peripheral retinal quadrants (collective P = 0.24). These results suggest that the posterior region of the human retina is susceptible to lipid peroxidation and that age is a factor.

Age Factors↗

PubMed1992

Lipid peroxidation in rod outer segments. Role of hydroxyl radical and lipid hydroperoxides.

Lipid peroxidation of rod outer segment (ROS) membranes has been implicated in the pathogenesis of numerous ocular disease processes. The hydroxyl radical might be involved in initiating the reaction. An in vitro system was developed to study lipid peroxidation of the ROS and the role of the hydroxyl radical. Bovine ROS were suspended in various concentrations of ferrous sulfate, incubated for 10 min at 37 degrees C, treated with diethylenetriamine pentaacetic acid to chelate the iron, and subjected to a thiobarbituric acid assay for malondialdehyde. A predictable increase in lipid peroxidation occurred in the presence of Fe+2. No effect was seen in the presence of Fe+3. Adding hydrogen peroxide, which would form the hydroxyl radical by reacting with Fe+2, had no effect at low concentrations. At higher concentrations, lipid peroxidation was inhibited, presumably from the oxidation of Fe+2 to Fe+3. Ethanol, a known hydroxyl radical scavenger, had no inhibitory effect in concentrations up to 0.50 mol/l. Conversely, cumene hydroperoxide and linoleic acid hydroperoxide, which form hydrophobic radicals, stimulated lipid peroxidation in the presence of Fe+2. These findings suggest that, under these experimental conditions, the hydroxyl radical is not an initiator of lipid peroxidation in ROS. They provide evidence that endogenous lipid radicals may initiate the reaction.

Animals↗

PubMed1991

Basic fibroblast growth factor promotes in vivo cerebral angiogenesis in chronic forebrain ischemia.

This experiment was designed to determine if intraventricular administration of basic fibroblast growth factor (bFGF) could promote cerebral angiogenesis in a model of mild chronic forebrain ischemia. Wistar rats underwent bilateral carotid artery ligation. Animals received intraventricular injections of bFGF every 4 days for 28 days. Basic fibroblast growth factor caused a significant dose-dependent increase in capillary density compared to ischemic controls in all regions examined. These results support the hypothesis that chronic intraventricular administration of bFGF induces in vivo cerebral angiogenesis.

Animals↗

PubMed1991

Corticotropin releasing factor antagonist reduces ischemic hippocampal neuronal injury.

This experiment tested the hypothesis that corticotropin-releasing factor (CRF) contributes to hippocampal ischemic injury. The antagonist to CRF (alpha-CRF) was administered intraventricularly 15 min prior to 10 min of transient forebrain ischemia in the Wistar rat. alpha-CRF demonstrated a neuroprotective effect in a dose-dependent manner most notable in CA1. There was also an increase in postischemic EEG recovery. It is postulated that CRF contributes to hippocampal ischemic injury through increased neuronal activity.

Animals↗

PubMed1991

Rat retina has an active and stable ubiquitin-protein conjugating system.

We describe here the presence of ubiquitin and its conjugation system in the rat retina. Retinal homogenates and supernatants conjugate [125I]human ubiquitin with either endogenous or exogenous proteins. The conjugating activity is relatively stable over time, requires ATP, and has a pH optimum of approximately 8. The most prominent [125I]ubiquitin conjugates formed are larger than 130 kDa. Several other minor conjugates are also formed between the molecular weights 17 and 75 kDa. The endogenous levels of free and conjugated forms of ubiquitin have been determined in the rat retina. More than 50% of retinal ubiquitin is covalently bound to target proteins. In addition, activities responsible for the ATP-dependent degradation and disassembly of both endogenous and exogenous ubiquitin conjugates have been detected in vitro. These results provide evidence that the retina contains active and stable ubiquitin-conjugating enzymes that recognize retinal proteins and have ATP-dependent proteolytic activity.

Adenosine Triphosphate↗

PubMed1991

Plasma lipid abnormalities in the miniature poodle with progressive rod-cone degeneration.

The miniature poodle with progressive rod-cone degeneration (prcd) is a model for human retinitis pigmentosa (RP). Since previous studies from several laboratories have shown abnormalities in plasma lipids in human RP, we examined the plasma lipids of prcd-affected animals. Fasting blood was drawn on three separate occasions from affected and control miniature poodles and on one occasion from normal Irish setters and those affected with a different inherited retinal degeneration (rod-cone dysplasia). Plasma phospholipids from prcd-affected animals had significantly lower levels of docosahexaenoic acid (22:6 omega 3) and cholesterol, compared to control miniature poodles. No differences were observed in plasma levels of phospholipids, vitamin E, or vitamin A, and no lipid differences were found between control and affected Irish setters. The ratios of 22:5 omega 3 to 22:6 omega 3 and of 22:4 omega 6 to 22:5 omega 6 were significantly elevated in prcd-affected poodles compared to controls. Since the conversion of 22:5 omega 3 to 22:6 omega 3 and of 22:4 omega 6 to 22:5 omega 6 is catalysed by a delta 4-desaturase, these results are consistent with a defect in desaturase activity in the prcd-affected poodle.

Animals↗

PubMed1991

Metabolism of linolenic acid and docosahexaenoic acid in rat retinas and rod outer segments.

Docosahexaenoic acid (22:6 omega 3) is uniquely enriched in photoreceptor outer segment phospholipids, comprising up to one-half of the fatty acids of phosphatidylethanolamine and phosphatidylserine. The current study was designed to investigate the incorporation of 22:6 omega 3 into outer segment phospholipids over 12 days and to determine whether the retina contained the enzymes necessary for elongation and desaturation of the major dietary precursor of 22:6 omega 3, the essential fatty acid linolenic acid (18:3 omega 3). Sprague-Dawley rats were injected intravitreally with [14C]22:6 omega 3 or [14C]18:3 omega 3 and kept in cyclic light (12 hr light/12 hr dark) for 2 hr to 12 days. Phospholipids from rod outer segments and the remaining retinal debris were separated by two-dimensional thin-layer chromatography. [14C]22:6 omega 3 radioactivity was initially highest in phosphatidylcholine and rapidly decreased from 45% of total phospholipid labeling at 2 hr to 26% by 1 and 3 days in ROS, while phosphatidylethanolamine labeling increased from 49 to 68% by 3 days and phosphatidylserine labeling increased from 3 to 14% over 12 days. Phenacyl derivatives of total fatty acids were separated by HPLC. A substantial conversion of [14C]18:3 to [14C]20:5, [14C]22:5 and [14C]22:6 was noted after 1 days, with increasing conversion to [14C]22:6 over the 12-day period. When only one eye was injected with [14C]18:3 omega 3, negligible radioactive fatty acids were detected in the contralateral eye from 1 to 12 days post-injection demonstrating that conversion of 18:3 to 22:6 occurred primarily within the injected eye. All enzymes for elongation and desaturation of 18:3 to 22:6 appear to be present in the eye. However, the conversion of 22:5 to 22:6 by delta-4 desaturase is evidently rate-limiting and may affect phospholipid replacement during photoreceptor outer segment renewal if this pathway proves to be essential for the supply of 22:6 during disk membrane formation.

Animals↗