Development and characterisation of a glutamate-sensitive motor neurone cell line

Modification of the growth conditions of NSC-34 mouse neuroblastoma x motor neurone cells by serum depletion promotes the expression of functional glutamate receptors as the cells mature into a form that bears the phenotypic characterisation of motor neurones. Immunocytochemical studies demonstrated the presence of the glutamate receptor proteins NMDAR1, NMDAR2A/B, GluR1, GluR2, GluR2/3, GluR4, GluR6/7, and KA2. Toxicity assays using cell counting techniques demonstrated a mild but significant cell death (approximately 30%, p < 0.01) following a 24-h exposure to 1 mM glutamate that could be prevented by the presence of the glutamate receptor antagonists (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (10 microM) and 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulphonamide (1 microM). As an indication of glutamate receptor functional activity a novel approach was used to detect the production of free radicals following stimulation with glutamate receptor agonists. The release of superoxide free radicals was detected using a micro-electrochemical sensor following addition of glutamate receptor agonists to the cell bathing solution. Alterations in intracellular calcium concentrations were examined using fura-2 imaging. Exposure of the differentiated NSC-34 cells to glutamate leads to an increase in intracellular calcium concentrations that is prevented by the presence of glutamate receptor antagonists. The motor neurone origin of these cells makes them particularly useful for investigating the potential role of glutamatergic toxicity in motor neurone degeneration.     

Response decrement in an auditory neurone of the locust

An identified interneurone (SA3) of the suboesophageal ganglion has an axon in both circumoesophageal connectives ascending to the brain. This is a novel morphology for a cell in the auditory system of the locust. The neurone is also novel for its physiological responses to sound in that it displays side-dependent response decrement (habituation). Responses to a tone directed at one ear decrement without affecting responses from the other ear. The responses on the decremented side recover when the opposite ear is stimulated. The decrement is the result of a diminishing amplitude of the compound excitatory post-synaptic potential but no inhibition is seen. Response decrement does not occur if the stimulus frequency is varied.     

Changes in metatarsal robusticity following experimental surgery

When a marginal metatarsal (M1 or M5) is amputated in neonatal rats, robusticity increases in all other metatarsals in both sexes. The robusticity increase is greatest in the metatarsal adjacent to the operation and declines in the direction away from it. The tripod structure of the rat's foot with its maximum robusticity of the marginal metatarsals is retained. When both marginal metatarsals are removed, this tripod arrangement is less clearly retained. When robusticity of all remaining metatarsals is averaged in a total robusticity quotient, total robusticity increase is greater in M1 than in M5 removal and greatest when both are removed. Total increase is greater in females than in males. Metatarsal robusticity increases result from retardation of longitudinal growth and increase of weight. The fact that some robusticity increases coincide with lesser length but not with increased weight suggests that in the process of robusticity increase longitudinal growth retardation precedes that of weight increase. Lesser length and greater weight of a long bone occurs also when hypofunction is produced in neonatal rats. However, in hypofunction the thickness of the compacta is increased leading to an obliteration of the medullary canal, while in the experimental metatarsals of this study, submitted to greater weight-bearing stresses, this canal is never reduced or obliterated.     

Co-administration of IGF-I and glycosaminoglycans greatly delays motor neurone disease and affects IGF-I expression in the wobbler mouse: a long-term study

The study on wobbler mouse has shown that the combined treatment with low doses of glycosaminoglycans (GAGs) and insulin-like growth factor-I (IGF-I) fully prevented motor neurone death and forelimb impairment up to 9-12 weeks of a mouse's life. The effect was accompanied by the prevention of the early hypertrophy of wobbler neurones, an effect likely due to the promotion of neuronal survival. At the 18th week, wobbler mice treated with IGF-I + GAGs still showed significantly improved forelimb function, reduced muscle atrophy and a higher number of cervical motor neurones. IGF-I alone and GAGs alone were active up to the 3rd week of treatment; thereafter the beneficial effects of single treatments decreased drastically. GAGs and IGF-I treatments also affected IGF-I plasma and muscle levels. In wobbler mice there was a progressive reduction in IGF-I plasma levels that was prevented by IGF-I or GAGs alone and greatly increased, even above heterozygote levels, by the combination treatment. Such a powerful increase was correlated by a small enhancement in insulin-like growth factor binding protein-3 (IGFBP-3) plasma levels, while treatment with IGF-I alone affected very significantly both IGFBP-1 and IGFBP-3. Co-treatment also prevented the decrease in IGF-I content observed in vehicle-treated wobbler mice forelimb muscles.     

Changes of brain gangliosides in chicken and mice during heterothermic development

Developmental profiles of brain gangliosides of chicken and mice were correlated to thermo-biological parameters. In parallel with an increasing body temperature and resistance to cold stress (within the first 11 postnatal days) the brain gangliosides change to a less polar pattern, indicated by a higher proportion of oligosialogangliosides.     

Changes in brain metallothionein and Zinc during development in transgenic mice

The developmental alterations in metallothionein (MT) proteins and zinc (Zn) were investigated in brains of two transgenic strains of mice. MT protein was measured by a cadmium binding assay and Zn by atomic absorption spectrophotometry. MT proteins were expressed at birth (day 1) both in MT-I overexpressing transgenic mouse (MT-I*) and MT-null (expressing only brain specific isoform, MT-III) transgenic mouse. MT proteins level (mainly MT-I) in MT-I* was 16.1 Μ-g/g at birth, and thereafter increased with age to a maximal adult level of 55.3 Μg/g (day 60). Zn level in MT-I* also increased from 8.43 Μg/g (day 1) to 20.7 Μg/g (day 60) with age. MT protein (MT-III) in MT-null mouse was 9.71 Μg/g at birth and remained relatively unchanged during development. Zn level in MT-null mouse at birth was 9.46 Μg/g and also remained unchanged during development. The similar alterations in MT isoforms and Zn in brain during development suggest that MT isoforms may act as a Zn binding protein.     

Changes in cytokinin activity during flower development in Cosmos sulphureus Cav

Endogenous cytokinin activity was determined in the flowers of Cosmos sulphureus Cav. from bud emergence to full bloom using the soybean callus bioassay. Cytokinin activity was low early in flower development but increased prior to full bloom. In Sephadex LH-20 column chromatography of flower extracts, the cytokinins present co-eluted with zeatin, zeatin riboside and glucoside cytokinin. While the former two predominated prior to full bloom, cytokinin glucoside activity appeared to be at a maximum at full bloom. The possible relevance of these findings is discussed in relation to flower development.     

Changes of the caecal villi during postnatal development in rats

Summary The villi of the caecal mucosa in postnatal rats were studied using both scanning electron and light microscopy.On the day of birth, numerous villi of various sizes and shapes were present on the caecal mucosa. After the 5th day, the villi decreased very rapidly in length and in number. A strong constriction was observed at the basal region of the caecal villi. During postnatal days 5 9 the villi probably separated and disappeared from the caecal mucosa. No villi were observed in rats that were over 10 days of age.     

Steroid control of neuron and muscle development during the metamorphosis of an insect

Insect metamorphosis is controlled by a small ensemble of developmental hormones including a class of steroids--the ecdysteroids. In the tobacco hornworm, Manduca sexta, the progression from the larval to pupal to adult stages is controlled by the relative blood titers of ecdysteroids and juvenile hormone (JH). The cellular events in the nervous and muscular systems which accompany metamorphosis resemble those of embryonic development, but they occur in an animal which is larger and experimentally more tractable than an embryo. In this paper we review the role of ecdysteroids in directing the metamorphosis of the nervous and muscular systems in Manduca, and how JH modifies the cellular responses to the steroids. In particular, we describe how these hormones control muscle degeneration, changes in the structure and function of identified neurons, and programmed neuron death. One general finding is that interactions between cells (e.g., neurons and their target muscles) are not involved in their hormonal responses, but rather the hormones act independently and in parallel at the different sites. Another key finding is that the critical periods and hormonal requirements for the commitment to a particular differentiative pathway, and the phenotypic expression of that pathway, can differ, and are therefore experimentally separable. Finally, we find that the significance of a hormonal signal (e.g., a rise in blood ecdysteroids) is interpreted differently depending upon the previous history of hormone exposure of a neuron or muscle. This progressive change in the interpretation of hormonal signals is a major mechanism by which a limited number of hormones can orchestrate a complicated phenomenon such as metamorphosis.     

The relationship between sensory stimulation and gross motor behaviour during the postnatal development in the rat

The behavioural development in the rat was studied from day 1 to day 12 under two stimulus conditions, one involving weak and the other strong stimulation. Depending upon the stimulus applied, different developmental schedules were obtained. Strong stimulation elicited complex behaviour patterns at an earlier age than did weak or moderate stimulation.     

Strength of forelimb lateralization predicts motor errors in an insect

Lateralized behaviours are widespread in both vertebrates and invertebrates, suggesting that lateralization is advantageous. Yet evidence demonstrating proximate or ultimate advantages remains scarce, particularly in invertebrates or in species with individual-level lateralization. Desert locusts (Schistocerca gregaria) are biased in the forelimb they use to perform targeted reaching across a gap. The forelimb and strength of this bias differed among individuals, indicative of individual-level lateralization. Here we show that strongly biased locusts perform better during gap-crossing, making fewer errors with their preferred forelimb. The number of targeting errors locusts make negatively correlates with the strength of forelimb lateralization. This provides evidence that stronger lateralization confers an advantage in terms of improved motor control in an invertebrate with individual-level lateralization.     

Changes in photosynthetic characteristics during leaf development in apple

A comprehensive developmental survey of leaf area, chlorophyll, photosynthetic rate, leaf resistance, transpiration ratio, CO₂ compensation point and photorespiration was conducted in apple. The largest changes in each of the photosynthetic characteristics studied took place during the earliest stages of leaf development, coinciding with the period of greatest leaf expansion and chlorophyll synthesis. During early development, photosynthesis increased 5-fold, reaching a maximum rate of 40 mg CO₂ dm^-2 hr^-1 at a leaf plastochron index (LPI) of 10. During this same period, leaf resistance, transpiration ratio, CO₂ compensation point and mesophyll resistance decreased, while carboxylation efficiency increased. Two especially interesting aspects of the data discussed are simultaneous changes that occur at a LPI of 10 and 12 in all of the photosynthetic characteristics examined and an apparent decrease in photorespiration as leaves age. From our results it is clear that stage of leaf development is an important factor affecting the rate of photosynthesis and photorespiration.Scientific Paper No. 5687, College of Agriculture, Washington State University, Pullman. This work is supported by the National Science Foundation Grant 80-10958 and the Columbia River Orchards Foundation.     

Motor neurone responses during a postural reflex in solitarious and gregarious desert locusts

Desert locusts show extreme phenotypic plasticity and can change reversibly between two phases that differ radically in morphology, physiology and behaviour. Solitarious locusts are cryptic in appearance and behaviour, walking slowly with the body held close to the ground. Gregarious locusts are conspicuous in appearance and much more active, walking rapidly with the body held well above the ground. During walking, the excursion of the femoro-tibial (F-T) joint of the hind leg is smaller in solitarious locusts, and the joint is kept more flexed throughout an entire step. Under open loop conditions, the slow extensor tibiae (SETi) motor neurone of solitarious locusts shows strong tonic activity that increases at more extended F-T angles. SETi of gregarious locusts by contrast showed little tonic activity. Simulated flexion of the F-T joint elicits resistance reflexes in SETi in both phases, but regardless of the initial and final position of the leg, the spiking rate of SETi during these reflexes was twice as great in solitarious compared to gregarious locusts. This increased sensory-motor gain in the neuronal networks controlling postural reflexes in solitarious locusts may be linked to the occurrence of pronounced behavioural catalepsy in this phase similar to other cryptic insects such as stick insects.     

Changes in bromodeoxyuridine labeling index during radiation treatment of an experimental tumor

Cell proliferation kinetics in a spontaneous mouse fibrosarcoma (FSaII) growing in C3H mice has been studied by in vivo pulse labeling of cells synthesizing DNA with bromodeoxyuridine (BrdUrd). A monoclonal antibody to BrdUrd and flow cytometry were used to quantify these cells. Labeling indices (LI) were measured before and after radiation. Unirradiated 10-mm tumors had a mean LI of 17.5%. After a single dose of 20 Gy there was depression of LI after 1 day followed by a rapid increase to greater than control values after 5 days. Analysis performed after five fractions showed that LI was dependent on the dose per fraction and interval between fractions. After 5 and 7 Gy/fraction LI remained similar to control values during daily fractionation but was significantly depressed after twice daily fractionation. With doses greater than 10 Gy/fraction there was marked depression of LI using both fractionation schedules. These changes in LI correlated well with changes in tumor volume after radiation. Tumors were also biopsied after 5 fractions of a 20-fraction course to see if LI would predict for tumor control. LIs of greater than or equal to 10% were associated with lack of tumor control at 90 days while all controlled tumors had a significant depression of LI. Changes in LI after radiation were a reasonable indication of the amount of repopulation occurring and might be useful in selecting patients for altered fractionation schedules.     

整合昆虫发育生物学和果蝇遗传学来研究昆虫发育与变态

成熟动物(昆虫)个体大小主要由生长持续时间和生长速度2个因素所决定。蜕皮激素和保幼激素协同调控昆虫发育变态,并决定昆虫生长持续时间;胰岛素、营养和细胞接触抑制等生长死亡信号及其传导途径控制细胞分裂、长大、分化、死亡,并最终决定昆虫的生长速度。最近研究成果表明,蜕皮激素信号和胰岛素信号相互影响,对昆虫个体大小起决定性的作用;脂肪体和营养代谢把这2条信号传导途径整合起来。科学家将会整合昆虫发育生物学和果蝇遗传学,抓住生长持续时间和生长速率2个关键因素,并以营养代谢和脂肪体为切入点来研究昆虫的发育变态。     

Changes in heat shock protein synthesis and heat sensitivity during mouse thymocyte development

Heat shock protein synthesis was examined in mouse thymocytes at three stages of development: early embryonic thymocytes, which are CD4^?CD8^?, adult thymocytes, which are primarily CD4⁺CD8⁺, and mature spleen T cells, which are CD4⁺CD8^? or CD4^?CD8⁺. After either a 41°C or 42°C heat shock, the synthesis of the maior heat-inducible protein (hsp68) was elevated during the first hour of recovery but then decreased abruptly in thymocytes from adult mice. In contrast, the synthesis of hsp68 continued for up to 4 h after heating embryonic mouse thymocytes or mature spleen T cells. The more rapid termination ofthe heat shock response in the adult thymocytes was not the result of eitherless heat damage or more rapid repair since the recovery of general protein synthesis was more severely delayed in these cells. As well, the double positive CD4⁺CD8⁺ cells were more sensitive to hyperthermia than either the double negative CD4^?CD8^? or single positive CD4⁺CD8^? or CD4^?CD8⁺ cells. Exposure of fetal thymus organ cultures to elevated temperature revealed that the double negative thymocytes were able to survive and differentiate normally following a heat shock treatment that was lethal for the double positive thymocytes. Exposure of thymocytes from adult mice to elevated temperatures induced apoptotic cell death. This was evident by the cleavage of DNA into oligonucleosome-sized fragments. Quantitation of the extent of DNA fragmentation and the number of apoptotic cells by flow cytometry demonstrated that the extent of apoptotic cell death was related to the severity of the heat stress. Double positive (CD4+CD8+) thymocytes are selected on the basis of their T-cell antigen receptor (TCR). Most of these cells are negatively selected and die within the thymus by an active process of cell deletion known as apoptosis. Restricting hsp synthesis in response to stress might be essential during developmental processes in which cell maturation is likely to result in death rather than functional differentiation. © 1993Wiley-Liss, Inc.     

Decreases in motor unit firing rate during sustained maximal-effort contractions in young and older adults

Previous studies have suggested that older adults may be more resistant to muscular fatigue than young adults. We sought to determine whether motor unit firing rate might be a factor that determines the response to fatiguing exercise in young and older subjects. Motor unit recordings and muscular forces were obtained from the tibialis anterior (TA) muscle of 11 young and 8 older individuals. Maximal voluntary force was first measured during maximal-effort dorsiflexion contractions. Each subject then performed a series of 15 maximal isometric contractions, with each contraction lasting 30 s. A 10-s rest period separated the fatiguing contractions. As a result of the fatiguing exercise, both subject groups demonstrated a significant loss in maximal force. The force decline was less in the older adults (20.4%) than in the young adults (33.8%). As expected, prior to muscle fatigue, maximal firing rates in the TA muscle were greater in the young (28.1 ± 5.8 imp/s) than in the older adults (22.3 ± 4.8 imp/s). The decrease in motor unit firing rate with fatigue was also greater in the young adults (34.9%), than in the older adults (22.0%). These results suggest that the greater fatigue-resistance exhibited by older individuals might be explained by the fact that the decline in motor unit firing rate during fatigue is greater in young persons than it is in older adults.     

Changes in jasmonic acid concentration during early development of apple fruit

Apple fruits ( Malus domestica Borkh.) were harvested from 24 to 136 days after full bloom (DAFB) and endogenous jasmonic acid was analyzed by GC-MS. There were two isomers of jasmonic acid in apple fruit with a ratio of 37:63 ( cis:trans ). The cis:trans ratio remained relatively constant throughout this period of fruit development. The endogenous jasmonic acid concentration was 138 ng g⁻¹ fresh weight 24 DAFB and decreased as fruit developed. Changes in jasmonic acid concentration were coincident with those of respiration, ethylene production, and anthocyanin accumulation in patterns consistent with the reported responses to exogenous jasmonates. Possible roles for jasmonic acid during early fruit development are discussed.     

Changes in the gonadotropic function of the rat hypophysis during development of experimental alcoholism

Luteinizing hormone (LH) and prolactin blood plasma levels and LH level in the pituitary of alcoholic male rats were studied Alcoholic rats (heavy drinkers) have revealed hyperprolactinemia and inadequate LH secretions. The possible role of gonadotropins in the development of hypogonadism in alcoholic rats is discussed.