Hippocampal damage after injection of kainic acid into the rat entorhinal cortex

This study analyses the anticipatory postural adjustments during the gait initiation process in children aged 2.5, 4, 6 and 8 years. In adults, anticipation during gait initiation includes a shift in the centre of foot pressure (CP) both backwards and towards the stepping foot. Backward displacement and the duration of the anticipation phase covary with the gait progression velocity reached by the subject at the end of the first step. In the present study, the children walked on a force plate that allowed us to calculate the acceleration of the centre of mass and the displacements of the CP. The results showed three main characteristics of the development of anticipatory behaviour: (1) The occurrence of anticipatory displacements of the CP increased progressively with age. Systematic backward anticipation was found for all children except one of the youngest, whereas the lateral displacement was systematically observed later, in the 6-year group; (2) the amplitude of the spatial parameters showed a significant increase with age; (3) contrary to the adult, the amplitude of the backward shift did not covary with the forthcoming velocity in the youngest groups. This covariation became significant at 6 years and remained significant at 8 years. The results showed that even if anticipatory behaviour was present in 2.5-year-old children it is only later that the child is able of more accurate tuning of feedforward control, probably due to better control of the overall postural adjustments.     

Dopamine receptors status after unilateral nigral 6-OHDA lesion. Autoradiographic and in situ hybridization study in the rat brain

The physiological effects of dopamine (DA) are mediated by several distinct receptor subtypes. The effects of unilateral nigral 6-hydroxydopamine (6-OHDA) lesions on DA receptors were investigated by receptor autoradiography using the D1 selective ligand [3H]SCH 23390 as well as the D2 ligand [3H]spiroperidol. mRNA distribution was studied by in situ hybridization. Lesioned rats were sacrificed at different time intervals. Receptor binding studies were performed on tissue sections using selective ligands. [35S]UTP labeled RNA probes were prepared from the different cDNA (D1, D2, D3) and used for in situ hybridization. A specific loss of receptor binding sites and mRNA hybridization was found in the lesioned substantia nigra pars compacta (SNc) at all times examined. Receptor binding studies revealed a different time-dependent increase in both D1 and D2 receptors. In situ hybridization showed that only D2 receptor mRNA increased in the caudate-putamen (CPu) of the lesioned side 15 d after 6-OHDA. No changes were observed in D1 and D3 receptor mRNA during the entire time-course.     

Neurotrophins and their receptors in the pigeon caecal tonsil. An immunohistochemical study

Neurotrophins are growth factors which bind to signal-transducing receptors called Trk proteins. The neurotrophins and their receptor proteins are present in the mammalian and avian lymphoid organs, thus suggesting that these factors could act upon cells of the immune system. Nevertheless, little is known about the cellular distribution of neurotrophins and their receptor proteins in avian lymphoid tissues. In this study we use immunohistochemistry to detect the cellular localisation of neurotrophins and their receptor proteins in the pigeon caecal tonsil, used as a model for avian secondary lymphoid organs. Rabbit polyclonal antibodies against neurotrophins (nerve growth factor -NGF-, brain-derived neurotrophic factor -BDNF- and neurotrophin -3 NT-3-) and against specific epitopes of TrkA, TrkB and TrkC proteins were used. Cytokeratins, vimentin, S-100 protein and chromogranin A were studied in parallel to identify cells which seemed to express neurotrophins and Trk proteins. TrkA-like protein was seen in the intestinal epithelium, whereas TrkB-like and TrkC-like proteins was found in cells which we identified as dendritic cells and macrophages. BDNF-like and NT-3-like reactivity was localised in intestinal epithelial cells, especially endocrine cells. Present results add further evidence to the presumptive immune role of neurotrophins and their receptors and the possible functions of these peptides in the caecal tonsil are discussed.     

Regulation of progesterone receptor messenger ribonucleic acid in the rat medial preoptic nucleus by estrogenic and antiestrogenic compounds: an in situ hybridization study

Progesterone receptor (PR) messenger RNA (mRNA) is concentrated in neurons of the preoptic area and other regions of the rat hypothalamus where it is colocalized with the estrogen receptor and regulated by changes in the steroid hormonal milieu. To date, little is known about the regulation of PR mRNA by estrogens and whether antiestrogenic compounds are capable of modulating its expression. The present studies used in situ hybridization to ascertain the time course of PR mRNA regulation in the medial preoptic nucleus by 17beta-estradiol, determine the effective dose required to elicit a response, and compare the efficacy of 17beta-estradiol with a variety of estrogenic or antiestrogenic compounds. The first series of studies revealed that the treatment of ovariectomized rats with 17beta-estradiol resulted in an increase in PR expression within 2 h, after which it remained elevated until 10 h postinjection and then returned to baseline levels. When ovariectomized rats were injected with 25-1000 ng/kg of 17beta-estradiol and euthanized 6 h later, a dose-dependent increase in the level of PR mRNA was observed, with a maximal response at 1000 ng/kg and an EC50 of 93.5 ng/kg. Subsequent studies evaluated the efficacy of a variety of estrogenic and antiestrogenic compounds in the rat preoptic nucleus. 17Beta-estradiol, diethylstilbestrol, and 17alpha-estradiol all significantly increased the level of PR mRNA, although the degree of induction varied with each compound. The injection of tamoxifen, raloxifene, toremifene, droloxifene, clomiphene, GW 5638, or ICI 182,780 had no significant estrogenic effect on PR gene expression at the dose evaluated. In contrast, when tamoxifen or raloxifene, but not ICI 182,780, was administered in the antagonist mode, a significant dose-related decrease in the estradiol-induced level of PR mRNA was seen in the preoptic area. The results of these studies clearly demonstrate that PR mRNA expression in the rat preoptic area is rapidly stimulated by a small dose of 17beta-estradiol. Moreover, the present report has also shown that the estrogenic nature of compounds such as tamoxifen, raloxifene, toremifene, droloxifene, clomiphene, and GW 5638 cannot be predicted by their activity in peripheral tissues. Together, the results of these studies provide important information about the central activity of estrogens and provide evidence for their tissue-specifc actions in the rat.     

Neurotrophins and their receptors in the tench retina during optic nerve regeneration

To understand the role of neurotrophins in the visual system, we investigated the distribution of both neurotrophins and their receptors within the retina of a fish that has the capacity to spontaneously regenerate its optic nerve axons after lesion. Intact retinas and retinas from tench, whose optic nerve had been crushed, were analyzed by immunohistochemistry and in situ hybridization. Trk receptors were mainly immunolocalized in cells of the inner nuclear and ganglion cell layers, a distribution coincident with that of their mRNAs. Nerve growth factor (NGF) immunoreactivity was detected exclusively in Müller cell processes, and brain-derived neurotrophic factor (BDNF) was found in both neuronal bodies and Müller cell processes. Neurotrophin-3 (NT-3) was detected in most of the cell nuclei, and neurotrophin-4/5 (NT-4/5) was localized in fibers and in a few cells in the inner retina. An increase in both TrkA protein and mRNA was detected during axonal regeneration within the retinal ganglion cell layer, reaching a maximum 30 days postcrush and returning to normal levels by day 90, when optic nerve regeneration is almost completed in this fish. None of the other neurotrophins and receptors showed appreciable changes. The heterogeneous distribution patterns of neurotrophins and their receptors in fish retina, their differences from the distribution observed in other species, and the TrkA changes after optic nerve crush suggest an important role for these molecules in the normal physiology of the fish retina and during the regeneration process.     

Regionally specific induction of BDNF and truncated trkB.T1 receptors in the hippocampal formation after intraseptal injection of kainic acid

The septo-hippocampal cholinergic and GABAergic systems were lesioned with single unilateral injections of kainic acid (KA) into the septum to further characterize the role of these afferents in the regulation of hippocampal brain-derived neurotrophic factor (BDNF) expression. Nearly all cells expressing choline acetyltransferase, trkA or glutamic acid decarboxylase mRNA disappeared in the medial septum 7 days after the neurotoxin administration. The lesion resulted in a complete loss of CA3 pyramidal cells, and robust increases in BDNF mRNA levels in hippocampal granular dentate cells and in the amygdala. There were rapid transient increases of BDNF mRNA levels in the hippocampal formation and cortex. In addition, we found a strong induction of truncated trkB.T1 mRNA receptors in the stratum radiatum and stratum oriens of the CA3 subfield. The prolonged induction of BDNF mRNA levels suggests an important role of this neurotrophin, possibly mediated by truncated trkB receptors, in the regulation of hippocampal plasticity following injury.     

Distribution of striatin, a newly identified calmodulin-binding protein in the rat brain: an in situ hybridization and immunocytochemical study

The worm-like chain model has often been employed to describe the average conformation of long, intrinsically straight polymer molecules, including DNA. The present study extends the applicability of the worm-like chain model to polymers containing bends or sections of different flexibility. Several cases have been explicitly considered: (i) polymers with a single bend; (ii) polymers with multiple coplanar bends; (iii) polymers with two non-coplanar bends; and (iv) polymers comprised of sections with different persistence lengths. Expressions describing the average conformation of such polymers in terms of the mean-square end-to-end distance have been derived for each case. For cases (i) and (iv), expressions for the projection of the end-to-end vector onto the initial orientation of the chain are presented. The expressions derived here have been used to investigate DNA molecules with sequence-induced bending (A-tracts). Mean-square end-to-end distance values determined from a large number of A-tract containing DNA molecules visualized by scanning force microscopy resulted in an average bend angle of 13.5 degrees per A-tract. A similar study was performed to characterize the flexibility of double-strandedDNA molecules containing a single-stranded region. Analysis of their mean-square end-to-end distance yielded a persistence length of 1.3 nm for single-stranded DNA.     

Expression of neurotrophins and their receptors in the developing and adult rat adrenal gland

We have studied the postnatal expression of neurotrophins, their cognate high-affinity trk receptors and the low-affinity NGF receptor (p75LNGFR) in the rat adrenal gland using RT-PCR. Neurotrophin mRNAs were detectable during the whole postnatal period. Strongest signals were obtained for BDNF and NT4/5. Expression of trkA, trkB, trkC and p75LNGFR was found at all ages studied. Signals for trkA were highest in the adult adrenal medulla, whereas signals for p75LNGFR were highest in the adult adrenal cortex. Cur data suggest still largely enigmatic roles for neurotrophins in functions of the adrenal medulla and possibly also the cortex.     

Changes in NMDAR2 subunit mRNA levels during pentobarbital tolerance/withdrawal in the rat brain: an in situ hybridization study

BACKGROUND: Urban academic medical centers provide care for large populations of vulnerable older adults. These patients often suffer a disproportionate share of chronic illnesses, disabilities, and social stressors that may increase health care costs. OBJECTIVE: To describe the distribution and content of total healthcare costs accrued over a 4-year period by a community of older adults cared for in an urban academic healthcare system and to describe high-cost patients and utilization patterns. DESIGN: A cohort study. SETTING: A tax-supported public healthcare system consisting of a 450-bed hospital and seven community-based ambulatory care centers. PATIENTS: 12,581 patients aged 60 years and older who had at least two ambulatory visits and/or one hospitalization within the healthcare system from 1993 through 1995. MEASUREMENTS: Patient demographic and clinical characteristics, hospital and ambulatory utilization rates, and all healthcare costs accrued from 1993 through 1996 were determined. Costs were estimated from the perspective of the healthcare system using cost to charge ratios. MAIN RESULTS: The mean patient age was 70 years, 60% were women, 44% were Black, and 83% were covered by Medicare and/or Medicaid. Nearly 25% of patients were obese, 15.8% had a history of smoking, and 15.5% had evidence of malnutrition. The mean number of ambulatory visits per year was 4.3 (+/-7.2), and 38.1% of patients had been hospitalized one or more times. Within the 4-year window, 24.1% of patients had missed five or more appointments with their primary care physicians, 32.7% of patients had five or more unscheduled clinic visits, and 12.5% had five or more emergency room visits. Total health care costs for 4 years for this cohort of older adults was $125.2 million dollars, with per capita annual mean costs of $3893. Expenditures associated with hospitalizations accounted for 63.6% of healthcare costs. Total inpatient and outpatient costs for the 38% of patients hospitalized at least once accounted for 85.3% of all health care expenditures. Patients who died in the hospital did not accrue significantly greater costs than patients who died out of the hospital. Simulations of a random 5% adverse selection of high-cost patients among two capitated systems resulted in cost shifts of $11.1 million. Recorded smoking history, obesity, and low serum albumin were significantly associated with excess costs. CONCLUSIONS: Healthcare costs are concentrated in a significant minority of older adults. Costs accrued in conjunction with hospital stays dominate healthcare expenditures for this cohort of older adults. However, most older adults (83%) have one or fewer hospital episodes in a 4-year period. Although patients who died accrued greater healthcare costs, these costs were not higher when the death occurred in the hospital. Self-care behaviors are an important target for interventions to reduce costs.     

Expression of acetylcholinesterase messenger RNA in human brain: an in situ hybridization study

The distribution of messenger RNA coding for acetylcholinesterase was studied in human post mortem brain and rhesus monkey by in situ hybridization histochemistry and compared to the distribution of acetylcholinesterase activity. Acetylcholinesterase messenger RNA had--similar to acetylcholinesterase enzymatic activity--a widespread distribution in human bain. Acetylcholinesterase messenger RNA positive cells corresponded to perikarya rich in acetylcholinesterase activity in most but not all regions. Examples for mismatches included the inferior olive and human cerebellar cortex. The presence of hybridization signals in cerebral cortex and an enrichment in layer III and V of most isocortical areas confirmed that perikaryal acetylcholinesterase in cerebral cortex is of postsynaptic origin and not derived from cholinergic projections. In striatum the expression of high levels of acetylcholinesterase messenger RNA was restricted to a small population of large striatal neurons. In addition, low levels of expression were found in most medium sized striatal neurons. Cholinergic neurons tended to express high levels of acetylcholinesterase messenger RNA whereas in cholinoceptive neurons the levels were moderate to low. However, some noncholinergic neurons like dopaminergic cells in substantia nigra, noradrenergic cells in locus coeruleus, serotoninergic cells in raphé dorsalis, GABAergic cells in thalamic reticular nucleus, granular cells in cerebellar cortex and pontine relay neurons expressed levels comparable to cholinergic neurons in basal forebrain. It is suggested that neurons expressing high levels of acetylcholinesterase messenger RNA may synthesize acetylcholinesterase for axonal transport whereas neurons with an expression of acetylcholinesterase confined to somatodendritic regions tend to contain lower levels of acetylcholinesterase messenger RNA.     

NMDA-NR1 receptor subunit mRNA expression in rat brain after 6-OH-dopamine induced lesions: a non-isotopic in situ hybridization study

Antisense digoxigenin-labeled deoxyoligonucleotides probes and non-isotopic in situ hybridization (HIS) techniques have been used to explore the NMDA-NR1 receptor subunit mRNA distribution in different brain areas of rats which had their dopaminergic nigrostriatal pathway previously lesioned with intracerebral administration of 6-OH-dopamine (6-OH-DA). Intense and significant hybridization signals for NR1 mRNA were found in dentate gyrus and regions CA1-CA2-CA3 of the hippocampus, in layers II-III and V-VI of the cerebral cortex, and in the cerebellum of sham-treated rats. Basal ganglia structures such as the striatum exhibited few NR1 mRNA hybridization signals as compared to the hippocampus and cerebral cortex. In contrast, both zona compacta and reticulata of substantia nigra (SN) showed a reduced number of cells with nevertheless intense NR1 mRNA HIS signals. The NR1 mRNA distribution in the brain was affected in a brain regional selective manner by 6-OH-DA induced lesions of DA neuronal systems. A striking increase in NR1 mRNA HIS signals was observed in both striata after unilateral lesioning with 6-OH-DA. Instead, in SN compacta but not in reticulata, a moderate but significant bilateral reduction of NR1 mRNA was observed after unilateral 6-OH-DA injection. No significant changes in NR1 mRNA were detected in cerebral cortex and other brain regions after 6-OH-DA treatment. These studies, and others reported in the literature, support the view that extensive lesions of nigrostriatal DA-containing neurons in the brain may trigger compensatory or adaptative responses in basal ganglia structures such as striatum and substantia nigra which involve glutamateric neurons and the genic expression of NMDA receptors.     

Expression of trkB and trkC mRNAs by adult midbrain dopamine neurons: a double-label in situ hybridization study

The documented trophic actions of the neurotrophins brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4/5 (NT-4/5) upon ventral mesencephalic dopamine neurons in vitro and in vivo are presumed to be mediated through interactions with their high-affinity receptors TrkB (for BDNF and NT-4/5) and TrkC (for NT-3). Although both neurotrophin receptor mRNAs have been detected within the rat ventral midbrain, their specific association with mesencephalic dopaminergic cell bodies remains to be elucidated. The present study was performed to determine the precise organization of trkB and trkC mRNAs within rat ventral midbrain and to discern whether the neurotrophin receptor mRNAs are expressed specifically by dopaminergic neurons. In situ hybridization with isotopically labeled cRNA probes showed that trkB and trkC mRNAs were expressed in all mesencephalic dopamine cell groups, including all subdivisions of the substantia nigra and ventral tegmental area, and in the retrorubral field, rostral and caudal linear raphe nuclei, interfascicular nucleus, and supramammillary region. Combined isotopic/nonisotopic double-labeling in situ hybridization demonstrated that virtually all of the tyrosine hydroxylase (the catecholamine biosynthetic enzyme) mRNA-containing neurons in the ventral midbrain also expressed trkB or trkC mRNAs. Additional perikarya within these regions expressed the neurotrophin receptor mRNAs but were not dopaminergic. The present results demonstrate that essentially all mesencephalic dopaminergic neurons synthesize the neurotrophin receptors TrkB and TrkC and thus exhibit the capacity to respond directly to BDNF and NT-3 in the adult midbrain in vivo. Moreover, because BDNF and NT-3 are produced locally by subpopulations of the dopaminergic cells, the present data support the notion that the neurotrophins can influence the dopaminergic neurons through autocrine or paracrine mechanisms.     

Intrathecal injection of lysine acetylsalicylic acid in the rat: a neurotoxicological study

Lysine acetylsalicylic acid has been reported to induce analgesic effects in humans after intrathecal (i.t.) injection. Before conducting further studies in humans with this drug, it is important to evaluate potential toxicological effects on the spinal cord in animals. In the present study the effects of chronic intrathecal administration of provocative doses of lysine acetylsalicylic acid (L-ASA) on the rat spinal cord were evaluated using light and electron microscopy and a quantitative morphometric method. We also investigated the effects of single doses of the drug on the spinal cord blood flow (SCBF) using the laser-Doppler flowmetry technique. No histopathological changes or differences in number or density of neuronal cells could be seen after chronic administration of L-ASA as compared to controls. The SCBF decreased immediately after i.t. injection of a large dose (4 mg) of L-ASA and returned to predrug levels within 10 min. At the end of the experiment metabolic acidosis was detected, indicating a systemic effect of acetylsalicylic acid. It is concluded that no neurotoxic effects on the spinal cord were seen after chronic i.t. injection of L-ASA. From a neurotoxicological point of view, our findings do not contraindicate the spinal use of L-ASA in humans.     

Differential expression of c-fos mRNA in rat prefrontal cortex, striatum, N. accumbens and lateral septum after typical and atypical antipsychotics: an in situ hybridization study

The regional difference in the expression of c-fos mRNA induced by typical and atypical antipsychotics was determined in prefrontal cortex, striatum, N. accumbens and lateral septum in rats by in situ hybridization. Two typical antipsychotics, haloperidol (2 mg/kg) and fluphenazine (2 mg/kg), and three atypical antipsychotics, (-)sulpiride (100 mg/kg), clozapine (20 mg/kg) and OPC-14597 (40 mg/kg), were used. Brains were fixed with 4% paraformaldehyde 45 min after drug administration (i.p.). Brain sections of 30 microns-thickness were made in a cryostat and hybridized with 35S-labelled for c-fos oligonucleotide probe. These sections were apposed to X-ray films and the autoradiograms were semi-quantitatively analysed by computer-assisted densitometry. All antipsychotics used increased c-fos mRNA expression in N. accumbens shell, a region of the forebrain associated with limbic systems. On the other hand, two typical antipsychotics (haloperidol and fluphenazine) that cause a high incidence of acute motor side effects increased the expression of c-fos mRNA in the dorsolateral striatum, an extrapyramidal region primarily involved in motor control. Only clozapine induced c-fos mRNA in the medial prefrontal cortex and lateral septum. These results strongly suggest that the shell region of N. accumbens may be a common site of therapeutic action of antipsychotics.     

Opioid receptor gene expression in the rat brain during ontogeny, with special reference to the mesostriatal system: an in situ hybridization study

Arachidonic acid metabolites are potent modulators in physiology and mediators in pathophysiology of airways. They play important role in allergic diseases. There are two main sources of eicosanoids found in nasal and bronchial lavages: airway epithelial cells and influx cells. Authors described spectra of eicosanoids produced by epithelial cells in vitro and compare them with in vivo findings. The review of similarities and differences between arachidonic acid metabolism in human upper and lower airways is also included.     

Neurotrophins prevent death and differentially affect tyrosine hydroxylase of adult rat nigrostriatal neurons in vivo

Brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4 (NT-4) promote survival of mesencephalic dopaminergic neurons in vitro and affect normal and damaged ones in vivo. Here, these neurotrophins had markedly different potencies to prevent the death of axotomized nigrostriatal dopaminergic neurons when infused close to the rostral end of the nigral nucleus of adult rats (NT-4 > BDNF > NT-3; nerve growth factor or NGF without effect). With a high dose of BDNF (30 micrograms/day) complete protection was achieved in the rostral but not caudal nigral regions, consistent with its poor diffusion characteristics in brain tissue. Measurements of tyrosine hydroxylase immunoreactivity suggest that BDNF and NT-4 (presumably through their TrkB receptor) reduce the synthesis of this rate-limiting enzyme for dopamine synthesis in rescued as well as in normal neurons. In sharp contrast, survival-promoting doses of NT-3 (presumably through its TrkC receptor) maintained normal levels of tyrosine hydroxylase immunoreactivity in the rescued nigrostriatal neurons. These results suggest that for these adult central nervous system neurons, some neurotrophic factors are predominantly involved in facilitating cell survival, whereas others are more involved in regulating neurotransmitter function.     

Bone marrow one step fixation-decalcification in Lowy FMA solution: an immunohistological and in situ hybridization study

The immunoreactivity of paraffin embedded bone marrow biopsies (BMB) was studied following a one step 20-hour-fixation-decalcification in Lowy formalin mercuric chlorid acid solution which permits excellent histological stainings. Antibodies reactive with myeloid, megakaryocytic, erythroid cells, T and B lymphocytes, mastocytes and metastatic cells were compared. Nearly all antibodies working on paraffin sections were demonstrated on Lowy FMA fixed BMB. Special care was taken to define an optimal working dilution. Trypsinization was not necessary. A slide microwave pre-treatment appeared essential before testing CD20 L26, CD8, CD3, CD34, MB1 Kappa and Lambda antibodies. It was suitable for UCHL1, LN2, CD30 antibodies. The same fixative allowed an m RNA Kappa or Lambda in myeloma and EBER 1 EBV RNAs in HIV lymphoma visualization by in situ hybridization. The safety handling of the toxic mercuric chloride component is discussed.