Mauro Bindoni

Change in neurotrophins and their receptor mRNAs in the rat forebrain after status epilepticus induced by pilocarpine

We studied the effects of status epilepticus (SE) induced by lithium chloride/pilocarpine treatment on gene expression of neurotrophins of the nerve growth factor (NGF) family and of their high‐affinity receptors of the tyrosine protein kinase (trk) family in the forebrain. Using in situ hybridization (ISH), we demonstrated an early (3 h after treatment) increase in brain‐derived neurotrophic factor (BDNF) and trkB mRNA expression in the dentate gyrus, amygdala, and piriform cortex, as well as widespread increases in the cerebral cortex. NGF mRNA, but not the mRNA of its receptor trkA, was increased in the dentate gyrus. In contrast, 12 h after treatment, neurotrophin‐3 (NT‐3) decreased, and its receptor trkC mRNA increased. There was no change in NT‐4 mRNA levels. All changes were blocked by pretreatment with scopolamine, a muscarinic antagonist. The noncompetitive N‐methyl‐aspartate (NMDA) antagonist ketamine blocked NGF, BDNF, and trkB mRNA increases in the hippocampus and cerebral cortex, but not in the amygdala and piriform cortex. In contrast, ketamine did not affect NT‐3 and trkC changes. These results provide a complete description of changes in mRNA levels of neurotrophins and their receptors in the forebrain after SE and supply additional data supporting the view that neurotrophin gene expression is related to abnormal neuronal activity.

Increased expression of trkB and trkC messenger RNAs in the rat forebrain after focal mechanical injury.

Tyrosine protein kinases trkA, trkB and trkC are signal transduction receptors for a family of neurotrophic factors known as the neurotrophins. Here we report on changes in the expression of messenger RNAs for trkA, trkB and trkC in the brain following an injury caused by insertion of a 30-gauge needle into adult rat hippocampus or neocortex. Quantitative in situ hybridization revealed no change in the level of trkA messenger RNAs in any brain region following this insult. In contrast, increased levels of trkB messenger RNA compared to untreated animals were seen in the granule cell layer of the dentate gyrus ipsilateral to the injury already 30 min after the injury. The increase reached maximal levels (four-fold) between 2 and 4 h, but returned to control levels 8 h after the injury. No change was seen in the contralateral dentate gyrus. The levels of trkC messenger RNA increased in the same brain regions as trkB messenger RNA, though with a delayed response, reaching a maximal increase of 3.3-fold 4 h after the injury. As for trkB messenger RNA, the level of trkC messenger RNA then tapered off and reached control levels 8 h after the injury. However, 4 h after the injury, a 1.7-fold increase of trkB and trkC messenger RNAs were seen in the ipsilateral piriform cortex. The increases of trkB and trkC messenger RNAs were confirmed using a nuclease protection assay. Increases of both trkB and trkC messenger RNAs were also seen in the piriform cortex, but not in the hippocampus, following needle insertion into the neocortex. Pretreatment of the animals with the non-competitive N-methyl-D-aspartate antagonist ketamine completely prevented the increases of trkB and trkC messenger RNAs, suggesting that the brain injury caused a release of glutamate with subsequent activation of N-methyl-D-aspartate receptors. In contrast, the anticonvulsive drug diazepam, the muscarinic antagonist atropine and the calcium-channel antagonist nimodipine had no effect on the increases of trkB and trkC messenger RNAs. Combined with previous data on the expression of neurotrophin messenger RNAs following similar injuries, our results support the hypothesis that increased levels of neurotrophins and their receptors could protect against neuronal damage following a brain insult.

Interleukin 2 modifies the bioelectric activity of some neurosecretory nuclei in the rat hypothalamus

Introduction of 15-30 U of interleukin-2 (IL-2) into the 3rd ventricle of Wistar rats was followed by a marked and significant decrease in neuron discharge frequency in the ventromedial nucleus of hypothalamus and a marked increase in the supraoptic and paraventricular nuclei. The monoclonal antibody ART62 partly blocked these effects. The conventional anti-IL-2 receptor monoclonal antibody ART18 had only a non-significant influence on the effects of IL-2. Since the supraoptic and paraventricular nuclei secrete the antidiuretic hormone, their excitation may offer a partial explanation of the considerable water retention observed during IL-2 therapy against neoplasia.

Effects of ozone on some biological activities of cells in vitro

The aim of this work was to study thein vitro effect of ozone on the 70 kDa family of inducible heat shock proteins (HSPs 70). We also performed tests to investigate possible toxic effects of ozone at the different doses employed. In human haematic mononucleated cells ozone at doses up to 20 μg/ml had no toxic effects and induced biosynthesis of the HSPs70. Biosynthesis of these proteins was greater at 40 μg/ml. In murine macrophages testing with tetrazolium salt (MTT), neutral red, and 2-deoxy-d-[1-3H]glucose uptake and study of the cell morphology showed a remarkable resistance or no toxic effects at a dose of 100 μg/ml also. Melanoma B16 murine cells assayed with the MTT test demonstrated less resistance to the toxic effects of ozone than normal cells. These results provide indications relevant to the problems of ozone therapy.

Characterization and partial purification of a substance in the pineal gland which inhibits cell multiplication in vitro

A substance which inhibits the in vitro multiplication of 3 cell strains, 37 RC, KB and NCTC clone 929, was characterized in the sheep pineal gland and partially purified using three successive chromatography techniques, respectively on Sephadex G-25, CM-cellulose and Biogel P 60 columns. The sheep cerebral cortex and liver also contain, but at much lower concentration than in the pineal, substance(s) that behave in different tests like the factor isolated from the pineal. The nature of the antimitotic substance from the pineal is as yet unknown. It is not destroyed by treatment with proteolytic enzymes, nor by boiling with 6 M HC1. It was established that it is different from the known antiblastic drugs such as Daunomycin and Methotrexate and from some active substances known to be present in the pineal, such as melatonin, secotonin and norepinephrine, which, in the same conditions, did not show any antimitotic activity. It was shown that when the concentration of the pineal factor in the culture medium was high enough (10 mug/ml), the inhibition of the KB cells multiplication was complete and irreversible. Microscopic examination of the treated cells showed that the morphological alteration was rapid (3--6 h) and deep, with shrinkage of both cytoplasm and nucleus, while with antiblastic drugs, morphological alteration proceeded slower (1--3 days) and appeared less pronounced.

Growth of Yoshida Ascites Tumor in the Rat after Radiofrequency Destruction of the Tuberoinfundibular Region of the Hypothalamus

The mitotic index of Yoshida ascites tumor cells was significantly higher in rats that underwent tuberoinfundibular destruction than those lesioned in other regions of the hypothalamus or in the cerebral hemispheres or in sham-operated animals. Survival was shorter in the rats lesioned in the tubero-infundibular and posterior hypothalamic regions. The rise in the mitotic index of Yoshida ascites tumor cells is consistent with the results of previous work showing significantly increased cell proliferation in the normal tissues of animals lesioned in the tuberoinfundibular region.

Effects of early destruction of the mouse arcuate nucleus by monosodium glutamate on age-dependent natural killer activity

The present study has examined the effect on age-dependent natural killer (NK) cell cytotoxic (NKCC) activity after the destruction of the arcuate nucleus (AR) of the hypothalamus in newborn mice with the neurotoxin monosodium glutamate (MSG). The NKCC activity was determined at different ages from the 18th day to the 54th week. This treatment resulted in depressed NKCC activity and in disappearance of its age-dependent pattern. A high correlation between NKCC depression and decreased numbers of large granular lymphocytes was also noted in the MSG-treated mice. The conclusion is that normal development of NKCC activity requires the maturation of AR functions.

Cytotoxic and cytostatic activity of copper(II) complexes. Importance of the speciation for the correct interpretation of the in vitro biological results.

The cytotoxicity of some copper(II) compounds against the mouse cancer cell line B16, murine L929, human KB cells, and fibroblasts was investigated. All the copper(II) systems tested were shown to have pronounced toxicity against transformed cells and a cytostatic effect against untransformed cells, i.e., human fibroblasts. A careful speciation of the actual in vitro conditions reveals that copper(II) is essentially present as mixed complexes formed with the amino acids of the culture medium, [Cu(glutamine)(histidine)] being the main species. It was found that the cytotoxic activity is related to the amount of copper(II) contained in the tested compounds.

Effects of ether lipid 1-O-octadecyl-2-methoxy-rac-glycero-3-phosphocholine and its analogs PAF and CPAF on the release of nitric oxide in primary cultures of rat astrocytes

Ether lipid 1-O-octadecyl-2-O-methoxy-rac-glicero-3-phosphocholine (ET-18-OCH3) is an immunomodulator with antineoplastic activity. Its analog compounds PAF and CPAF share some of its biological effects. In our experiments, even very small amounts of ET-18-OCH3 released a remarkable quantity of nitric oxide (NO) from rat astrocytes cultured in vitro. The NO biosynthesis was inhibited by pretreatment with the antagonist BN 50730. The effect of ET-18-OCH3 was greater than that of the LPS inducer. PAF did not produce NO, even at high doses, while the nonmetabolizable CPAF only induced a significant release of NO from 12 micrograms/ml onwards. These results demonstrate that ET-18-OCH3 is functionally active also in astrocyte cultures. Stimulation of NO biosynthesis is of a great value on account of its the known effect as a neurotransmitter, potentiator of immune defences and possible modulator of cerebral circulation.

Physiological and pathological influences of central nervous system on the immune system: A critical appraisal

There is considerable evidence of intricate links between central nervous and immune systems. This paper makes a critical assessment of these relationships. Recent experimental data on hypothalamic influences on Natural Killer activity in mice have been presented and the problems met with in defining causal nexuses discussed. Some information on immune reactivity in patients with mental disorders has briefly been reviewed.