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Biomedical subjects

I C Campbell

Publications and source records attributed to I C Campbell.

At least 19 recordsLinked to original sources
PubMed2000

A mathematical model for assessing changes in neurofilament protein levels in neurites and cell bodies of differentiating neuroblastoma cells.

A mathematical model which allows the calculation of the level of neurofilament protein in the cell body (x) and in the neurites (y) of differentiating SK-N-SH cells is presented. The model considers the changes in cell number (proliferating cells) and the number of cells with neurites (differentiating cells). It takes into account the fact that (i) when cells are cultured in differentiating conditions, an increase in cell number is initially observed and (ii) in a non-synchronized population of differentiating cells, the length of neurite extended by individual cells varies within the population. Total neurofilament protein levels in a population of cells were measured by enzyme-linked immunoabsorbant assay and application of the model to the data allowed values for x and y to be calculated. The validity of the model is supported by the fact that the predicted total neurofilament protein levels are highly correlated with the experimentally derived neurofilament protein levels. The model should be of use in temporal studies of cytoskeletal proteins involved in neuronal growth/differentiation and also in studies in which the system is a target of toxic insult.

Cell Differentiation↗
PubMed2000

Recombinant nef HIV-IIIB protein is toxic to human neurons in culture.

The expression of HIV-1 negative factor (nef) has been positively correlated with HIV disease progression [Z. Hanna, D.G. Kay, N. Rebai, A. Guimond, S. Jothy, P. Jocicoeur, Nef harbors a makor determinant of pathogenicity for an AIDS-like disease induced by HIV-1 in transgenic mice. Cell 95 (1998) 163-175]. Nef expression has been detected in HIV infected human brains with neuronal damage [A. Ranki, M. Nyberg, V. Ovod, M. Haltia, I. Elovaara, R. Raininko, H. Haapsalo, K. Krohn, Abundant expression of HIV Nef and Rev proteins in brain astrocytes in associated with dementia, AIDS 9(9) (1995) 1001-1008; Y. Saito, L.R. Sharer, M.G. Epstein, J. Michaels, M. Mintz, M. Londer, K. Golding, B.M. Blumberg, Overexpression of nef as a marker for restricted HIV-1 infection of astrocytes in postmorten paediatric central tissues, Neurology 14 (1994) 474-480]. It is postulated that nef may contribute to the neuronal damage observed in the brain of those with late HIV disease. To test this, the potential toxicity of recombinant nef (from HIV-1 IIIB) was compared to the neurotoxin human tumour necrosis alpha (TNFalpha) on human brain cells in culture. SK-N-SH neuroblastoma, primary human neurons and glial cells were exposed to recombinant nef or TNFalpha protein for 3 days or twice over 6 days. Cell viability was assessed by Trypan Blue, lactate dehydrogenase (LDH) release and MTT assays. Nuclear fragmentation was detected using the Hoechst Blue nuclear dye assay. Both nef and TNFalpha (100 ng/ml) caused a significant 30% reduction of SK-N-SH cell numbers after 3 days exposure (P=0. 001). At this time, exposure to nef caused evident fragmented nuclei in these cultures. Human neuronal cultures had a 32 and 33% decrease in cell number after 6 days exposure to either nef or TNFalpha, respectively (P<0.001). Furthermore, as previously shown [J. He, C.M. DeCastro, G.R. Vandenbark, J. Busciglio, D. Gabuzda, Astrocyte apoptosis induced by HIV-1 transactivation of the c-kit protoonocogene, Proc. Natl. Acad. Sci. 94 (1997) 3954-3959], a 3-day exposure to nef significantly reduced human glial cell number by 25% (P=0.001). Recombinant nef and TNFalpha compromise human neurons in culture. Thus, like other virotoxins, it is shown for the first time that nef may also contribute to neuronal damage that has been reported in dementia in late HIV disease.

Cell Nucleus↗
PubMed2000

Changes in neurite outgrowth but not in cell division induced by low EMF exposure: influence of field strength and culture conditions on responses in rat PC12 pheochromocytoma cells.

The effects of low electromagnetic field (EMF) exposure (4.5-15.8 microT, 50 Hz AC) on neurite outgrowth and cell division in rat PC12 pheochromocytoma cells were examined. The study involved two separate experimental series in which culture conditions during exposure to the magnetic fields differed. In series 1 (14 experiments in which culture conditions were not strongly conducive to cell differentiation [15% serum]), exposure to 4.5-8.25 microT for 23 h significantly inhibited neurite outgrowth by 21.5 +/- 1.3% (by Manova, p = 0.003). In contrast, in series 2 (12 experiments in which culture conditions promoted cellular differentiation [4% serum]), exposure to 4.35-8.25 microT for 23 h significantly stimulated neurite outgrowth by 16.9 +/- 1.1% (by Manova, p = 0.009). Thus, in both series, exposure to a narrow range of low EMF has significant, but opposite effects on neurite outgrowth. Exposure to higher fields, 8.25-12.5 microT (series 1) and 8.25-15.8 microT (series 2) had no significant effect on neurite outgrowth. These data, when considered with other reports, suggest that neuronal differentiation can be altered by low level EMF exposure. While this may not be detrimental, it merits further research. At present, the reasons for the significant changes in neurite outgrowth being confined to the same narrow field strength are unclear. As stated above, culture conditions in series 2 were more conducive to cell differentiation than those in series 1. This is reflected in the lower number of cells in control samples in series 2, at the end of the 23-h incubation, than in series 1 (- 16.9 +/- 1.7%, p = 0.003). As the same numbers were plated in both series, the medium used in series 1 allows more of the PC12 cells to divide; this is consistent with some cells reverting to a non-neuronal adrenal chromaffin phenotype [L. Greene, A. Tischler. Establishment of a noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor. Proc. Natl. Acad. Sci. U. S. A., 73 (1976) 2424-2426]. Exposure to both ranges of magnetic fields (4.35-8.25 and 8.25-15.8 microT) has no effect on cell division. Thus, there is no evidence in this study that there is a mitogenic effect arising from low EMF exposure.

Adrenal Gland Neoplasms↗
PubMed1999

Circadian regulation of prion protein messenger RNA in the rat forebrain: a widespread and synchronous rhythm.

Although the expression of the normal prion protein in the host is critical to the development of transmissible spongiform encephalopathies, the physiological role of this protein and the processes regulating its expression remain obscure. We now report that the messenger RNA for the prion protein is regulated in the rat brain in a marked circadian manner not only in the suprachiasmatic nuclei, the principal site for the generation of mammalian circadian rhythms, but also in other forebrain regions. The data show a remarkable consistency in the concurrence of a single peak of prion protein messenger RNA at each of the sites early in the animal's phase of increased locomotor activity; behavioural arousal does not, however, appear to affect this expression. We believe this to be the first study demonstrating that the expression of prion protein messenger RNA can change over a relatively short period in vivo. The results are discussed with reference to the range of recently discovered "clock-related" transcripts which also have widespread tissue expression; these include the messenger RNAs for D-box binding protein and thyroid embryonic factor, transcription factors which bind to the prion protein promoter.

Animals↗
PubMed1998

Phosmet induces up-regulation of surface levels of the cellular prion protein.

Chronic (2 day) exposure of human neuroblastoma cells to the organophosphate pesticide phosmet induced a marked concentration-dependent increase in the levels of PrP present on the cell surface as assessed by biotin labelling and immunoprecipitation. Levels of both phospholipase C (PIPLC)-releasable and non-releasable forms of PrP were increased on the plasma membrane. These increases appear to be due to post-transcriptional mechanisms, since PrP mRNA levels as assessed by Northern blotting were unaffected by phosmet treatment. These data raise the possibility that phosmet exposure could increase the susceptibility to the prion agent by altering the levels of accessible PrP.

Cell Membrane↗
PubMed1998

Variation in the expression of the mRNA for protein kinase C isoforms in the rat suprachiasmatic nuclei, caudate putamen and cerebral cortex.

Using in situ hybridization, we have examined mRNA expression for five isoforms of protein kinase C (PKC alpha, beta1, beta2, gamma and epsilon) in the rat suprachiasmatic nuclei (SCN) and other central site during the 24 h cycle. The signal for each of these isoforms shows a marked local density within the SCN. In the absence of photic cues, there are changes in the expression of the mRNAs for the four isoforms that are Ca2+-dependent (alpha, beta1, beta2 and gamma), but not for one of the Ca2+-independent PKCs (epsilon). PKC alpha mRNA exhibits a monophasic rhythm of expression in the SCN with a peak at early subjective night, circadian time (CT) 14. In contrast, the mRNAs for PKC beta1, beta2 and gamma show a biphasic rhythm in the SCN with peaks at early subjective day, CT 0, and early subjective night, CT 14. The four Ca2+-dependent isoforms may therefore subserve phase-related functions within the SCN at the onset of subjective night and, in the case of beta1, beta2 and gamma, also at the onset of subjective day. Variation in the mRNAs for PKC beta1 and gamma (but not for alpha, beta2 or epsilon) is also found in the caudate putamen and in the cingulate and parietal cortex; the biphasic pattern of expression for these mRNAs is precisely in phase with that observed in the SCN. The beta1 and gamma isoforms may therefore contribute to temporally regulated functions at sites outside the SCN. The present observations raise the possibility that receptor-mediated regulation of circadian functions is modulated or even gated by circadian changes in intracellular components that participate in distinct signal cascades.

Animals↗
PubMed1998

Neuroendocrine, appetitive and behavioural responses to d-fenfluramine in women recovered from anorexia nervosa.

BACKGROUND: Increased central serotonin (5-HT) function has been hypothesised to be a vulnerability trait in anorexia nervosa. METHODS: Eighteen women with a history of DSM-III-R anorexia nervosa and 18 female controls were examined. The subjects had recovered weight and menstrual function. A placebo-controlled d-fenfluramine test was used. Subjects ingested d-fenfluramine or placebo and after three hours were offered a 'free' meal. The amounts eaten were recorded and plasma cortisol and prolactin levels were measured. Questionnaires related to eating attitudes and behaviour, to personality, and to mood were administered. RESULTS: Unlike the control subjects, those recovered from anorexia nervosa did not show the expected appetite-suppressing responses to d-fenfluramine; their eating attitudes and behaviour were more restrained, 'negative' perfectionism was more pronounced, and post-meal plasma cortisol levels did not rise as expected. CONCLUSIONS: Our results do not suggest that increased central 5-HT function is a trait marker in anorexia nervosa, but dysregulation in part of the central 5-HT system may be a vulnerability factor. The flattened post-meal response to cortisol in the subjects who had recovered from anorexia nervosa suggests that their hypothalamic pituitary--adrenal axis may be altered and deserves further investigation.

Adult↗
PubMed1996

PKC in rat cortical synaptosomes: effects of depolarization.

PKC isotypes were studied in rat cortical synaptosomes. Resting synaptosomes, subfractionated into cytosolic and particulate (Triton X100-soluble and insoluble) fractions, containing PKC beta 1, gamma, delta and epsilon isotypes but very little PKC beta 2 or alpha. PKC delta and epsilon were evenly distributed in the cytosolic and particulate fractions; PKC beta 1 was mainly in the cytosol (70%) and PKC gamma was primarily particulate (80%). Triton X-100 extracted most of the PKC delta and epsilon from the particulate fraction but only half of the PKC gamma, indicating PKC gamma association with the cytoskeleton. Following KC1 treatment (30 mM), both the classical PKC isotypes beta 1 and gamma shifted from the cytosolic to the particulate fraction, with PKC beta 1 moving specifically to the detergent insoluble fraction whereas PKC gamma appeared to move to both. It is concluded that PKC beta 1, gamma, delta, and epsilon isotypes can occur presynaptically and suggested that PKC beta 1 and gamma are directly involved in synaptic transmission. Apparent mol. wt changes in translocating forms may be related to phosphorylation and subsynaptosomal location.

Animals↗
PubMed1996

The growth hormone response to apomorphine at 4 days postpartum in women with a history of major depression.

We have examined the responsiveness of dopamine sensitive neurones in the postpartum period in woman with a history of major depression who are at high risk of experiencing a recurrence of illness in the postpartum period. Fourteen women were assessed at 36 weeks of pregnancy and during the 3 months following delivery, using the Schedule for Affective Disorders and Schizophrenia, including its change version. They were not depressed at initial assessment. Five of the 14 women went on to experience a postpartum relapse (2 major depressive disorder, 2 generalised anxiety disorder, 1 panic disorder). On the fourth day postpartum, i.e., before relapse, the growth hormone response to the dopamine agonist apomorphine was measured as an index of the functional state of hypothalamic dopamine D2 receptors. Women who subsequently relapsed had a significantly greater growth hormone response to apomorphine than those who remained well. This was particularly marked in women with anxiety/panic. The development of increased sensitivity of hypothalamic dopamine D2 receptors in the postpartum period appears to predict the onset of depressive and anxiety disorders.

Anxiety Disorders↗
PubMed1996

Circadian changes of glutamate decarboxylase 65 and 67 mRNA in the rat suprachiasmatic nuclei.

Gaba has been implicated in the regulation of the circadian clock within the suprachiasmatic nuclei (SCN). In the present study in situ hybridization histochemistry was used to assess expression of mRNA for the two isoforms of the enzyme governing GABA synthesis, glutamic acid decarboxylase (GAD) 65 and 67, within the rat SCN during a 24 h cycle in constant darkness. GAD 65 mRNA exhibited a monophasic rhythm, with a peak at the beginning of the subjective day and a nadir early in the subjective night. In contrast, there was a biphasic variation in GAD 67 expression, with peaks at the beginning of the subjective day and early in the subjective night. No significant variation in the mRNAs for GAD 65 or GAD 67 was observed in the cingulate cortex. These results suggest that the clock intrinsic to the SCN controls the expression of the mRNAs encoding GAD 65 and GAD 67 in both a coordinated and a differential manner.

Animals↗
PubMed1996

Rapid identification of novel genes expressed in a circadian manner in rat suprachiasmatic nuclei.

The circadian clock in mammals is located in the suprachiasmatic nuclei (SCN). There is evidence that changes in gene expression are central to its mechanism. We are engaged in identifying the genes involved. The small size of the SCN, the large number of mammalian genes and the need to identify those differentially expressed over 24 h have required novel experimental procedures. mRNA differential display and an improved tissue micropunching method have been used to examine temporal changes in gene expression in the SCN of rats maintained in constant darkness. Several of the displayed cDNA species were found to be differentially expressed; they show no homology with published sequences. Riboprobes of these cDNA species were used for in situ hybridization. Emulsion-dipped sections confirmed that at least two of these differentially displayed mRNAs are expressed in a circadian manner.

Animals↗
PubMed1996

Circadian variation of EAAC1 glutamate transporter messenger RNA in the rat suprachiasmatic nuclei.

Using in situ hybridization, we examined temporal changes of the EAAC1 glutamate transporter mRNA within the suprachiasmatic nuclei (SCN) of rats in constant darkness. Film autoradiographs showed that the SCN and supraoptic nuclei (SON) contained a marked density of hybridization signal. Analysis of silver grains per cell in emulsion-dipped sections indicated that cellular expression of EAAC1 mRNA in the SCN was elevated during the latter part of the subjective night and at the beginning of the subjective day, with a peak at circadian time 23.1 as determined by cosinor analysis. The times at which EAAC1 mRNA is highest correspond to the time points at which extracellular glutamate, a neurotransmitter that putatively mediates photic entrainment, has been reported to be low within the SCN. The presence of EAAC1 mRNA in the SCN and SON may partially explain the resistance of these nuclei to glutamate receptor-mediated excitotoxins; furthermore, the raised level preceding subjective dawn in the SCN may ensure sub-toxic levels of extracellular glutamate at the onset of photic stimulation during the LD cycle. In contrast, cellular expression of EAAC1 mRNA in the cingulate cortex and reticular thalamus remained constant at all time points studied. These results suggest that there is circadian control of the EAAC1 mRNA by the clock intrinsic to the SCN.

Amino Acid Transport System X-AG↗
PubMed1995

Regulation of intracellular free calcium levels by the cellular prion protein.

A rat brain synaptosomal model was used to investigate the possible role of the cellular prion protein (PrP) in the regulation of intracellular free calcium levels ([Ca2+i). Treatment of synaptosomes with bacterially derived recombinant human PrP in the range 20-100 micrograms ml-1 resulted in dose-dependent elevations of [Ca2+]i. These increases were dependent on extracellular calcium and were inhibited by gadolinium chloride, a potent blocker of voltage-sensitive calcium channels. Conversely, when calcium channels were activated by synaptosomal depolarization, treatment with monoclonal antibody to PrP in the range 200-320 ng IgG ml-1 resulted in a dose-dependent reduction of [Ca2+i, which was blocked by competition with PrP preparations. These results indicate that PrP is associated with regulation of intracellular free calcium levels through an interaction with voltage-sensitive calcium channels.

Animals↗
PubMed1995

S-adenosyl-L-homocysteine and 5'-methylthioadenosine inhibit binding of [3H]flunitrazepam to rat brain membranes.

The effect of inhibitors of transmethylation reactions, S-adenosylhomocysteine and 5'-methylthioadenosine, on [3H]flunitrazepam-specific binding to the rat brain membranes has been investigated. Both S-adenosylhomocysteine and 5'-methylthioadenosine are able to inhibit binding with Ki values of 7.9 microM and 15.8 microM respectively. These compounds therefore may be candidate endogenous benzodiazepine-receptor ligands. In light of these and other data possible correlations between phospholipid methylation and gamma-aminobutyric acid (GABA) receptor function are discussed.

Animals↗
PubMed1995

Effects of sodium on PKC translocation; relationship to neurotransmitter release.

Our previous work using Na+ channel activators such tityustoxin (TsTX), indicated that local increases in Na+ modulate glutamate release from synaptosomes. We have now investigated the role of the Ca2+/phospholipid-dependent protein kinase (PKC) in mediating this effect. TsTX and KCl stimulate 'fast' glutamate release to the same extent but TsTX is more effective than KCl in enhancing the 'slow' phase of release. KCl greatly stimulates PKC translocation. However, TsTX inhibits basal and phorbol ester-induced translocation while the Na(+)-ionophore, gramicidin D, has no effect. Taken together, these data suggest TsTX mediated localized Na+ entry inhibits PKC translocation and that this effect may be associated with recruitment of vesicles to the readily releasable pool.

Acetylcholine↗
PubMed1995

Intestinal cellular immunity after primary rotavirus infection.

Infection of neonatal gnotobiotic lambs with a bovine strain of rotavirus was used to characterize the kinetics of the primary cellular intestinal immune response to this agent. At 2-3 days after infection virus was first detected in the faeces and increased numbers of CD45R+ cells were observed in peripheral blood. These cells persisted in significantly increased numbers in the circulation until 7-8 days after infection. At this time, virus was no longer detectable in the faeces. The increase in CD45R+ cells preceded the appearance of virus-neutralizing antibodies in the serum at 1 week after infection. Maximal antibody titres were reached 2 weeks after infection. Virus-primed cells were first observed 1 week after infection in the jejunal and ileal Peyer's patches, mesenteric lymph nodes and peripheral blood, and persisted in the mesenteric lymph nodes and jejunal Peyer's patches for a further 4 weeks. Analysis of lymphocyte surface antigens indicated that different sub-populations of lymphocytes were responding in the various lymphoid tissues; a majority of CD4+ cells was observed in the mesenteric lymph nodes, whereas B cells predominated in the ileal Peyer's patches.

Animals↗