Experimental protein malnutrition in primates--cytochemistry of the nervous system

Experimental protein malnutrition was induced in groups of young juvenile squirrel monkeys by feeding them ad libitum diets very low in protein content, whereas a diet containing 25% protein content was fed to the control animals. Detailed cytochemical studies have clearly shown the sensitivity of the nervous system to dietary abuse. The motor neurons of the spinal cord and the Purkinje cells of the cerebellum are very sensitive to protein deficiency, and the number of oligodendroglia cells increases sharply. Gallocyanin stained preparations from the malnourished animals show significant decrease in the amount of RNA in the Purkinje cells of cerebellum and the anterior horn cells of the spinal cord. The larger neurons show prominent chromatolysis with concomitant increase in the number of oligodendrocytes surrounding such nerve cells. The different layers of the cerebellar cortex of the malnourished animals, as well as some of the neurons of the spinal cord, showed decreased activity of succinate dehydrogenase and increased levels of thiamine pyrophosphatase and lactate dehydrogenase. Although ATPase activity appeared to remain unchanged quantitatively, it showed profound disturbance in its in situ localization, especially in the mitochondrial ATPase located in the cytoplasm. The enzyme activity in the nuclear membrane appeared unchanged. These studies emphasize the importance of studying in situ changes in the anatomically heterogeneous nervous tissue, rather than of working only with biochemical methods using homogenized material.     

Nucleolar changes in response to dietary protein malnutrition in the neurons of the motor cerebral cortex and cerebellum of squirrel moneky Saimiri sciureus.

Nucleolo-cytoplasmic relationships have been studied in healthy squirrel monkeys and those subjected to a known degree of protein malnutrition. In the latter group, thirty-two pregnant animals starting from 35 days of gestation and 24 young adult animals were given a diet containing 7.5% and 2.0% protein content, respectively, compared to a diet with 25% protein for the controls. The motor cortex and the cerebellum removed from neonates as well as young adult animals sacrificed after 9, 11, 13 and 15 weeks of feeding schedules were investigated. Four animals after 15 weeks of dietary protein deprivation were rehabilitated with a balanced diet over a year's period. Formaldehyde-fixed as well as fresh frozen tissues were used for the histological study and to employ histochemical techniques for the demonstration of lipids, carbohydrates, nucleic acids and enzymes of various metabolic cycles. As a result of protein malnutrition, the nucleolus in a majority of the neurons from the motor cortex and the Purkinje cells of the cerebellum undergoes a series of morphological and cytochemical transformations in response to cytoplasmic changes related to impaired protein metabolism. The greater the level of protein deprivation, the greater is the degree of cytoplasmic chromatolysis and more pronounced are the nucleolar transformation in terms of enlarged size, secretory activity and transfer of nucleolar material in the cytoplasm. The nucleolar buds located close to the periphery of the nuclear membrane and the nucleolar material in the cytoplasm show identical cytochemical nature except for the presence of DNA in the former. It appears that during migration through the nuclear membrane the nucleolar material loses its DNA component and only aggregates of ribosomes and protein pass into cytoplasm, which aid in the synthesis of specific proteins lost as a result of catabolic processes initiated by protein malnutrition. Most of the observed changes in the adult squirrel monkeys are reversed when they are rehabilitated with a balanced diet.     

Differential effects of protein malnutrition and ascorbic acid deficiency on histidine metabolism in the brains of infant nonhuman primates

Abstract: Male infant nonhuman primates (M. nemes-trina) born in captivity were used in the study. They were divided into three groups. The first group of three animals was fed a 20% casein diet and the second group of six monkeys received a 2.0% casein diet. The third group of four monkeys received a 20% casein diet totally devoid of ascorbic acid for 3.5 weeks before the diet was supplemented with ascorbic acid (20 mg/kg diet). All the diets were given to the animals in two daily rations of 100 g/animal. The monkeys fed a 2% casein diet failed to grow, and after about 3.5 months showed variable degrees of edema, hypoalbuminemia, evidence of psychomotor disturbance, depressed plasma levels of many essential amino acids, and other features consistent with the diagnosis of protein-energy malnutrition. Examination of the brains revealed significant alterations in the levels of histidine (+ 172%) and homocarnosine (+ 146%) in comparison with the control well-fed monkeys. Associated with the increase in brain histidine was a marked elevation of brain histamine level. Protein deficiency also led to poor brain retention of ascorbic acid but not to the same degree observed in the ascorbic acid-deficient animals. The latter group of animals, after receiving their diet for about 8 months, demonstrated a modest elevation in the plasma levels of most amino acids in comparison with controls. Ascorbic acid deficiency elicited a significant reduction (p < 0.01) in brain level of histidine, with hardly any change in homocarnosine level. In addition, vitamin C deficiency produced elevation of brain histamine level comparable to findings in the protein-energy-deficient monkeys. The results suggested that protein deficiency raised brain histamine level mainly through increased availability of the precursor amino acid histidine, while defective degradation might account for the increased brain level of this amine in ascorbic acid-deficient monkeys. Histamine has been proposed to have a predominantly depressant action on relevant neurons, and has also been shown to participate with other neuro-transmitters in influencing the function of the pituitary gland by regulating release of the hypothalamic hormones into the portal vessels. The relevance of the findings of marked increases in brain histamine in experimental protein and ascorbic acid deficiencies to the behavioral and extensive endocrinological alterations seen in human malnutrition deserves some intensive investigation.     

Cytochemistry of experimental protein malnutrition in primates: effect on the spinal cord of the squirrel monkey, Saimiri sciureus

Synopsis The localization of certain phosphatases, esterases and dehydrogenases in the spinal cord of healthy and severely protein malnourished squirrel monkeys were investigated histochemically. The latter were given drastically reduced levels of proteins in their diets for 15 weeks, and for the purpose of comparison several animals were sacrificed after 9, 11, 13 and 15 weeks on the feeding schedule. Cytochemical investigations were carried out on sections prepared from fresh-frozen spinal cord removed from the animal in the shortest possible time after an appropriate dose of nembutal anaesthesia. The distribution of thiamine pyrophosphatase, inosine diphosphatase, acid phosphatase and ATPase was found to be significantly altered under the impact of dietary abuse. The changes appear to be related to altered protein metabolism, energy transport and general slowing down of the metabolic reactions. The results obtained on phosphatases are reinforced by the distribution of enzymes of the Krebs cycle, pentose shunt and anaerobic metabolic pathways. Whereas the activity of the Krebs cycle enzymes is greatly reduced, the pentose shunt reacts to protein deprivation by increasing the level of activity of its enzymes. Similarly the activities of enzymes of the anaerobic pathway are also enhanced. The significance of these observations and the role of glial cells along with the neuron as a functional unit are discussed.     

Neurotoxicity of Glucocorticoids in the Primate Brain

Severe and prolonged physical and psychological stress is known to cause brain damage; long-term torture victims in prison bare later developed psychiatric disorders and cerebral cortical atrophy observed in CT scans (Jensen, Genefke, Hyldebrandt, Pedersen, Petersen, and Weile. 1982). In nonhuman primates, we observed degeneration and depletion of the hippocampal neurons in African green monkeys that had been severely abused by cagemates and died with complications of multiple gastric ulcers and adrenal cortical hyperplasia (Uno, Tarara, Else, Suleman and Sapolsky, 1989). In our previous studies the administration of dexamethasone (DEX) (5 mg/kg) to pregnant rhesus monkeys at 132 to 133 days of gestation induced degeneration and depletion of the hippocampal pyramidal and dentate granular neurons in the brains of 135-gestation-day fetuses, and these changes were retained in the brains of fetuses at near term, 165 days of gestation (Uno, Lohmiller, Thieme, Kemnitz, Engle, Roecker, and Farrell, 1990). We also found that implantation of a cortisol pellet in the vicinity of the hippocampus in adult vervet monkeys induced degeneration of the CA3 pyramidal neurons and their dendritic branches (Sapolsky, Uno, Rebert, and Finch, 1990). Thus, hippocampal pyramidal neurons containing a high concentration of glucocorticoid receptors appear to be highly vulnerable to either hypercortisolemia caused by severe stress or to exposure to exogenous glucocorticoids. To study the long-term postnatal sequelae of prenatal brain damage, eight rhesus monkeys were treated with either DEX (5 mg/kg), 5 animals, or vehicle, 3 animals, at 132 to 133 days of gestation. After natural birth, all animals lived with their mothers for 1 year. At 9 months of age, we found that DEX-treated animals had significantly high plasma cortisol at both base and post stress (isolation) levels compared to age-matched vehicle-treated animals. Magnetic resonance images (MRI) of the brain at 20 months of age showed an approximately 30% reduction in size and segmental volumes of the hippocampus in DEX-treated compared to vehicle-treated animals. Measurements of whole brain volume by MRI showed no significant differences between DEX and vehicle groups. Prenatal administration of a potent glucocorticoid (DEX) induced an irreversible deficiency of the hippocampal neurons and high plasma cortisol at the circadian baseline and post-stress levels in juvenile rhesus monkeys. These results suggest that the hippocampus mediates negative feedback of cortisol release; a lack or deficiency of the hippocampal neurons attenuates this feedback resulting in hypercortisolemia. The hippocampal deficiency in rhesus monkeys induced by prenatal administration of DEX appears to be a good model for neuroendocrinological dysfunctions and hippocampal development in human juveniles whose mothers were exposed to severe stress or received a high dose of glucocorticosteroids during pregnancy.     

EFFECTS OF DIFFERENT LEVELS OF DIETARY PROTEIN ON BRAIN GLUTAMATE DEHYDROGENASE AND DECARBOXYLASE IN YOUNG ALBINO RATS

Abstract— Studies were carried out to identify the minimum levels of protein (casein) needed in the diet in order to prevent or reverse the deficits in brain enzymes previously found with protein deficiency. Groups of weanling albino rats were fed diets containing variable amounts of protein (5, 8, 10, 15 or 20 per cent in experiment I, and 5, 6, 7, 8 or 20 per cent in experiment II) for 5 or 10 weeks. Deficits in brain wt and brain glutamate dehydrogenase and decarboxylase were found to be prevented by a diet containing 8 per cent or more of protein, although for optimum growth 15 per cent protein in the diet was found to be necessary. Groups of rats were fed a 5 or 20% protein diet for 10 weeks after which the 5% protein animals were either continued on the diet for another 10 weeks or changed to one containing 8, 10, 15 or 20% protein. The brain enzyme deficits found with the 5% protein diet were found to be fully reversed by feeding a 10% protein diet during rehabilitation.     

Changes in somatodendritic but not terminal dopamineregulation in aged rhesus monkeys

For these studies, young (8-9 years), middle-aged (14-17 years) and aged (23-28 years) rhesus monkeys were used as a model of normal aging in humans to investigate changes in dopamine (DA)-containing neurons in senescence. Aged monkeys exhibited significant age-related motoric declines as compared to the young animals. In vivo microdialysis studies showed that basal levels of the DA metabolites, homovanillic acid (HVA) and 3,4-dihydroxyphenylacetic acid (DOPAC) were diminished by 44% and 79%, respectively, in the substantia nigra (SN) of aged monkeys. In addition, d-amphetamine-evoked overflow of DA in the SN was diminished by 30% in the middle-aged animals and 67% in the aged monkeys. Post-mortem measures of DA and DA metabolites showed significant decreases in DA (20%), DOPAC (47%) and HVA (22%) levels in the putamen and a 25% decline in HVA tissue levels in the SN of the aged monkeys as compared to the young animals. Unbiased stereological cell counting of tyrosine hydroxylase (TH)-immunoreactive neurons in the SN showed a small (15-20%) but significant age-related decline in TH-positive neurons. In addition, there was a small (15-20%) but significant decline in TH-positive fiber density and TH-positive cell size. In comparison to the massive loss of DA neurons responsible for the movement dysfunctions seen in Parkinson's disease, pronounced functional changes in DA release in the SN and putamen may significantly contribute to the motoric dysfunctions characterizing normal aging in rhesus monkeys.     

Upregulation of anti-apoptotic factors in upper motor neurons after spinal cord injury in adult zebrafish

Unlike mammals, fish motor function can recover within 6–8 weeks after spinal cord injury (SCI). The motor function of zebrafish is regulated by dual control; the upper motor neurons of the brainstem and motor neurons of the spinal cord. In this study, we aimed to investigate the framework behind the regeneration of upper motor neurons in adult zebrafish after SCI. In particular, we investigated the cell survival of axotomized upper motor neurons and its molecular machinery in zebrafish brain. As representative nuclei of upper motor neurons, we retrogradely labeled neurons in the nucleus of medial longitudinal fasciculus (NMLF) and the intermediate reticular formation (IMRF) using a tracer injected into the lesion site of the spinal cord. Four to eight neurons in each thin sections of the area of NMLF and IMRF were successfully traced at least 1–15 days after SCI. TUNEL staining and BrdU labeling assay revealed that there was no apoptosis or cell proliferation in the axotomized neurons of the brainstem at various time points after SCI. In contrast, axotomized neurons labeled with a neurotracer showed increased expression of anti-apoptotic factors, such as Bcl-2 and phospho-Akt (p-Akt), at 1–6 days after SCI. Such a rapid increase of Bcl-2 and p-Akt protein levels after SCI was quantitatively confirmed by western blot analysis. These data strongly indicate that upper motor neurons in the NMLF and IMRF can survive and regrow their axons into the spinal cord through the rapid activation of anti-apoptotic molecules after SCI. The regrowing axons from upper motor neurons reached the lesion site at 10–15 days and then crossed at 4–6 weeks after SCI. These long-distance descending axons from originally axotomized neurons have a major role in restoration of motor function after SCI.     

Distribution of spinal sensory neurons supplying the adrenal gland in the pig

The present study was aimed at disclosing which spinal ganglia contribute to the innervation of the adrenal gland in the pig with special regard to the morphology and intraganglionic distribution of the primary sensory neurons within these ganglia. To this end, the animals were injected with a retrograde fluorescent neuronal tracer Fast Blue into the left adrenal gland during laparotomy. After a survival period of three weeks, labelled spinal sensory neurons were found in the ipsilateral dorsal root ganglia (approximately 99% of all retrogradely marked neurons). However, single adrenal gland-projecting perikarya were observed also in the contralateral dorsal root ganglia from Th3 to L3. The majority of the retrogradely labelled afferent neurons (above 65% of all Fast Blue-positive (FB+) perikarya) were located in two groups of spinal ganglia (at neuromeres Th6-7 and Th12-14, approximately 18% and 47% of neurons, respectively), forming two distinct multiganglionic centres of origin for this neural pathway. The morphometric evaluation of FB+ neurons revealed that the vast majority of them (approximately 81%) belonged to the class of small-sized perikarya (10-30 microm in diameter), while the medium-sized (diameter 30-80 microm) and large neurons comprised only up to 13% and 6.5% of adrenal gland-projecting neurons, respectively. Furthermore, the analysis of the intraganglionic distribution pattern of the retrogradely labelled cells revealed that the highest number of them was located in the medio-caudal domain of the dorsal root ganglia, irrespective of the neuromere studied. Thus, the present study has revealed sources and morphological characteristic of spinal afferent neurons supplying the porcine adrenal gland, simultaneously pointing out to the characteristic features of their inter- and intraganglionic distribution pattern.     

Effects of iron deficiency upon the antibody response to influenza virus in rats

The effects of severe and moderate iron deficiency upon the antibody response to influenza virus were investigated in rats. Three groups of weanling male Wistar rats were fed one of two iron-deficient diets (5 mg and 15 mg iron/kg diet) or a normal iron-containing diet (35 mg iron/kg diet). A group of individually pair-fed rats was introduced with the low iron-consuming rats. The effects of the diets upon various iron status parameters were followed during the 4th, 5th, 6th, and 7th week of diet. After 4 weeks of feeding different diets, an intraperitoneal injection of inactivated influenza virus A/New Jersey/76 was performed and a recall injection was done at 5 weeks. Primary and secondary antibody responses were assayed. Rats were sacrificed at 7 weeks of diet. After 4 weeks of feeding different diets, the rats fed the 5 mg iron/kg diet were severely anemic and rats fed 15 mg iron/kg diet were moderately iron-deficient, as shown by their iron status parameters. Growth was delayed in anemic and matched pair-fed rats. A primary antibody response was almost nonexistent in all groups. Secondary antibody titers were significantly weaker in anemic rats than in ad libitum controls, but were not different from those of pair-fed rats. This response was similar in moderately iron-deficient, ad libitum, and pair-fed rats. These results show that antibody synthesis in response to the influenza virus vaccine is preserved in moderate iron deficiency but is reduced in severe anemia. The reduction in energy consumption associated with severe iron deficiency in the rat could play a part in the altered humoral response.     

Immunohistochemical and in situ hybridization studies of choline acetyltransferase in large motor neurons of the human spinal cord

The localization of choline acetyltransferase (ChAT) protein and mRNA was investigated in large motor neurons of the lumbar spinal cord of 10 autopsied individuals without neurological diseases, by immunohistochemistry and in situ hybridization. In the immunohistochemistry using 20 serial tissue sections with a total thickness of 80 microm, about approximately 58-85% (average 67%) of the large motor neurons (30 microm and more in somal minimal diameter) in the ventral horn were stained with the anti-human ChAT antibody. In the positive neurons, most immunoreactive products were observed focally in the perikarya. Occasionally, the perikarya of some neurons were stained diffusely. In situ hybridization with a single 4 microm-thick tissue section showed that almost all large motor neurons had positive signals (approximately 93-100%, average 98%), which were distributed diffusely in the perikarya. The positivity rate in the in situ hybridization was higher than that in the immunohistochemistry for all 10 cases. These results indicate that ChAT mRNA is transcribed in almost all large motor neurons in the ventral horn of the human spinal cord, but ChAT protein cannot always be detected in the cytoplasm by immunohistochemistry.     

Influence of selenium deficiency on the acute cardiotoxicity of adriamycin in rats

The influence of selenium (Se) deficiency on the acute cardiotoxicity induced by the anticancer drug adriamycin (ADR) has been studied in rats by electrocardiography. Two categories were formed by feeding groups of rats a Se-supplemented and a Se-deficient diet. The supplemented animals were taken as normals. The two categories were treated with iv injections of saline solution containing ADR at doses of 0, 7.5, and 15 mg/kg body wt. The cardiac Se concentration and glutathione peroxidase (GSH-Px) activity in the Se-deficient groups were <2% lower than in the normals. The normal groups showed significant widening of the SaT and QaT durations when given 15 mg/kg ADR. The Se-deficient groups exhibited a dose-dependent widening of the SaT and QaT duration at 7.5 and 15 mg/kg and narrowing of the PQ duration at 15 mg/kg ADR. No heart rate or QRS duration changes were detected in both categories. Our results suggest that an imbalance of the antioxidant system is associated with Se deficiency and that Se plays a role in preventing the cardiac functional disorder attributable to oxygen free radical formation induced by ADR.     

Effect of serotonin on the formation of neurons producing gonadotropin-releasing hormone in Wistar rats

The effect of serotonin on the formation of neurons producing gonadotropin-releasing hormone (GnRH) during embryogenesis of Wistar rats was studied. The neurons producing GnRH were detected immunocytochemically on days 18 and 21 of embryonic development and on day 15 of postnatal development of rats with normal serotonin metabolism and rats in which the synthesis of serotonin was inhibited by p-chlorophenylalanine. The total number of GnRH neurons in serotonin deficiency was larger than in the case of its normal metabolism at all developmental stages studied. This is an indirect evidence for the inhibitory effect of serotonin on the formation of GnRH neurons. To confirm the morphogenetic effect of serotonin, we studied the rate of formation of GnRH neurons by injecting bromodeoxyuridine in the formation period of these neurons. It was found that serotonin deficiency had no effect on the time of formation of GnRH neurons: over 97% of neurons formed on days 11 to 15 of embryonic development both in the experimental and control groups. Note that, in serotonin deficiency, the maximum number of GnRH neurons formed one day later than in the normal state. Thus, serotonin inhibits the proliferation of GnRH neuron progenitor cells and thereby has a morphogenetic effect on the development of these neurons.     

Components of the translational machinery are associated with juvenile glycine receptors and are redistributed to the cytoskeleton upon aging and synaptic activity

The translation eukaryotic elongation factor 1alpha (eEF1A) is a monomeric GTPase involved in protein synthesis. In addition, this protein is thought to participate in other cellular functions such as actin bundling, cell cycle regulation, and apoptosis. Here we show that eEF1A is associated with the alpha2 subunit of the inhibitory glycine receptor in pulldown experiments with rat brain extracts. Moreover, additional proteins involved in translation like ribosomal S6 protein and p70 ribosomal S6 protein kinase as well as ERK1/2 and calcineurin were identified in the same pulldown approaches. Glycine receptor activation in spinal cord neurons cultured for 1 week resulted in an increased phosphorylation of ribosomal S6 protein. Immunocytochemistry showed that eEF1A and ribosomal S6 protein are localized in the soma, dendrites, and at synapses of cultured hippocampal and spinal cord neurons. Consistent with our biochemical data, immunoreactivities of both proteins were partially overlapping with glycine receptor immunoreactivity in cultured spinal cord and hippocampal neurons. After 5 weeks in culture, eEF1A immunoreactivity was redistributed to the cytoskeleton in about 45% of neurons. Interestingly, the degree of redistribution could be increased at earlier stages of in vitro differentiation by inhibition of either the ERK1/2 pathway or glycine receptors and simultaneous N-methyl-D-aspartate receptor activation. Our findings suggest a functional coupling of eEF1A with both inhibitory and excitatory receptors, possibly involving the ERK-signaling pathway.     

Gastric lipolysis in the developing rat. Ontogeny of the lipases active in the stomach

Two nutritional models, an essential fatty acid deficiency model and the feeding of saturated versus unsaturated fats, were used in a feeding study in order to assess the relationship between tissue fatty acid composition and the activities of some membrane-associated enzymes. Purified diets containing 7% hydrogenated coconut oil, 7% corn oil, 10% safflower oil or butter were fed to rats for a total of 49 weeks (1 week of pregnancy, 3 weeks of lactation and 45 weeks post-weaning). Tissue homogenates from submandibular salivary glands and kidneys were analyzed for fatty acid composition of total lipids and phospholipids. Changes in fatty acid patterns typical of essential fatty acid deficiency such as an increase in the levels of 16:1 and 18:1, a decrease in 18:2 and 20:4 and an accumulation of 20:3 omega 9 were observed in salivary glands and kidneys of rats fed the deficient diet. Tissues of rats fed 10% butter also showed fatty acid compositional changes which were somewhat similar to those in essential fatty acid deficiency, but to a lesser degree. The activities of ouabain-sensitive (Na+ + K+)-ATPase were higher in homogenates of salivary glands and kidneys of the deficient rats and those fed butter as compared with their controls. The results suggest a relationship between the double bond index of fatty acids as an indication of membrane lipid fluidity and allosteric modification of (Na+ + K+)-ATPase activity. However, other explanations for the observed changes in (Na+ + K+)-ATPase activity cannot be ruled out. There were no diet-related differences in the activities of gamma-glutamyltranspeptidase or 5'-nucleotidase.     

Fas and FasL Expression in the Spinal Cord Following Cord Hemisection in the Monkey

The changes of endogenous Fas/FasL in injured spinal cord, mostly in primates, are not well known. In this study, we investigated the temporal changes in the expression of Fas and FasL and explored their possible roles in the ventral horn of the spinal cord and associated precentral gyrus following T(11) spinal cord hemisection in the adult rhesus monkey. A significant functional improvement was seen with the time going on in monkeys subjected to cord hemisection. Apoptotic cells were also seen in the ventral horn of injured spinal cord with TUNEL staining, and a marked increase presents at 7 days post operation (dpo). Simultaneously, the number of Fas and FasL immunoreactive neurons in the spinal cords caudal and rostral to injury site and their intracellular optical density (OD) in the ipsilateral side of injury site at 7 dpo increased significantly more than that of control group and contralateral sides. This was followed by a decrease and returned to normal level at 60 dpo. No positive neurons were observed in precentral gyrus. The present results may provide some insights to understand the role of Fas/FasL in the spinal cord but not motor cortex with neuronal apoptosis and neuroplasticity in monkeys subjected to hemisection spinal cord injury.     

Dietary effects on brain fatty acid composition: the reversibility of n-3 fatty acid deficiency and turnover of docosahexaenoic acid in the brain, erythrocytes, and plasma of rhesus monkeys

Rhesus monkeys given pre- and postnatal diets deficient in n-3 essential fatty acids develop low levels of docosahexaenoic acid (22:6 n-3, DHA) in the cerebral cortex and retina and impaired visual function. This highly polyunsaturated fatty acid is an important component of retinal photoreceptors and brain synaptic membranes. To study the turnover of polyunsaturated fatty acids in the brain and the reversibility of n-3 fatty acid deficiency, we fed five deficient juvenile rhesus monkeys a fish oil diet rich in DHA and other n-3 fatty acids for up to 129 weeks. The results of serial biopsy samples of the cerebral cortex indicated that the changes of brain fatty acid composition began as early as 1 week after fish oil feeding and stabilized at 12 weeks. The DHA content of the phosphatidylethanolamine of the frontal cortex increased progressively from 3.9 +/- 1.2 to 28.4 +/- 1.7 percent of total fatty acids. The n-6 fatty acid, 22:5, abnormally high in the cerebral cortex of n-3 deficient monkeys, decreased reciprocally from 16.2 +/- 3.1 to 1.6 +/- 0.4%. The half-life (t 1/2) of DHA in brain phosphatidylethanolamine was estimated to be 21 days. The fatty acids of other phospholipids in the brain (phosphatidylcholine, -serine, and -inositol) showed similar changes. The DHA content of plasma and erythrocyte phospholipids also increased greatly, with estimated half-lives of 29 and 21 days, respectively. We conclude that monkey cerebral cortex with an abnormal fatty acid composition produced by dietary n-3 fatty acid deficiency has a remarkable capacity to change its fatty acid content after dietary fish oil, both to increase 22:6 n-3 and to decrease 22:5 n-6 fatty acids. The biochemical evidence of n-3 fatty acid deficiency was completely corrected. These data imply a greater lability of the fatty acids of the phospholipids of the cerebral cortex than has been hitherto appreciated.     

The effect of dietary protein on the mineral status of vervet monkeys with special reference to the impact of milk solids on calcium excretion

This study assessed the impact of Westernised and traditional African diets on mineral metabolism in general and calcium status in particular in vervet monkeys. Twelve adult male vervet monkeys (Cercopithecus aethiops), with an average weight of 5+/-0.58 kg each, were divided into two groups of six individuals each and fed traditional diets containing largely maize + legumes (17.4%) or Westernised diets containing milk solids (17.2%) as the source of high crude protein for 8 weeks. Blood was taken at 2-week intervals, the animals were weighed, while urine and stool samples were collected over 24 hours. The monkeys on the milk solids diet had diarrhoea for 6 weeks post-dietary intervention, and produced significantly greater quantities (P<0.02) of stool. These animals also produced significantly more urine (P<0.02). There was no difference in the degree of calciuresis of the two groups, but the monkeys on maize + legume proteins absorbed significantly more calcium during weeks two and six (P<0.04). Furthermore, both groups of monkeys showed a significant decline in plasma calcium levels over the experimental period (P<0.001). The diets had no effect on phosphate levels in the plasma or urine. However, both groups of animals absorbed less phosphate (P<0.09). There was an increasing loss of urinary magnesium (P = 0.03) in both groups, with the milk solids group showing lower plasma levels of this element (P = 0.09). However, the milk solids group lost less magnesium through the stool (P<0.03). In addition, the animals on milk solids showed significant natriuresis (P<0.05), while plasma sodium levels in both groups declined over time (P<0.03). Both diets induced a state of urinary potassium loss (P = 0.0003) and decrease in plasma potassium (P<0.0002). Urinary pH and plasma urea were unaffected by the diets, but the monkeys on maize + legumes excreted significantly less (P<0.001) urinary urea. This study indicates that the milk solids diet compromised mineral homeostasis by interfering with gut and renal functioning, while the traditional African diet did not induce these effects.     

Fos protein expression in the medulla oblongata and changes in size of spinal lateral horn neurons after 4-wk simulated weightlessness in rats

Responses of the neurons in medulla oblongata and C8-T1 spinal cord lateral horn of rats induced by simulated weightlessness were investigated using anti-Fos protein and anti-tyrosine hydroxylase (TH) double staining immunohistochemical methods, and Nissl-staining technique respectively. After four weeks of tail-suspension, many Fos-like positive neurons were localized in the medullary visceral zone (MVZ), predominantly in the nucleus of tractus solitarii and ventrolateral medulla, and some of them showed TH-like immunoreactivity. Sizes of the cell bodies of the lateral horn neurons in C8-T1 segment were significantly increased in 4-wk tail-suspended rats (P<0.05) as compared with that in controls. The results suggest that the neurons in MVZ and the spinal lateral horn may be involved in the adaptation of central cardiovascular regulation during weightlessness.     

Development of nitrogen and methane losses in the first eight weeks of lactation in Holstein cows subjected to deficiency of utilisable crude protein under restrictive feeding conditions

Low-protein diets are increasingly being used in dairy cow nutrition to minimise noxious nitrogen (N) emissions. However, at parturition, the lower milk yield at that time may mask deficiency in dietary utilisable crude protein (uCP; equivalent to metabolisable protein). Under restrictive feeding conditions, farmers would limit the feed allowance to match the lower measured milk yield, thereby exacerbating the deficiency. The consequences for N emission intensity per kg milk yield and methane emissions are unknown. In this study, two diets were fed to nine Holstein cows each from parturition onwards. One diet was complete and the other was calculated as 20% deficient in uCP. Feed allowance was always oriented towards the measured milk yield. In each of the first eight lactation weeks, intake and excretion were measured for 5 d. On the last 2 d of this period, methane emission was measured in respiration chambers. The statistical model included treatment, week and interaction as effects. The real levels of uCP and energy supply across the 8 weeks were 33% and 15% below requirements, respectively, in the Deficient cows. In addition, the Deficient cows consumed 18% less dry matter (caused by substantial refusals in week 1, where energy supply was according to requirements) and produced 25% less milk (26 vs. 34 kg/d). Cows in both groups used dietary N with similar efficiency for milk protein synthesis and excreted similar proportions of the N ingested via urine and faeces. This resulted in both treatments having similar N emission intensities per kg milk N and similar urinary N as a proportion of total excreta N, suggesting a similar potential for gaseous N emissions from the manure per kg of milk. The Deficient cows emitted 22% less methane overall but had similar methane yield and emission intensity to the Controls. In conclusion, a reduction in crude protein intake immediately after parturition does not reduce N emission per unit of milk when associated with uCP deficiency.