Interneuron-specific Ca2+ Responses Linked to Metabotropic -and lonotropic Glutamate Receptors in Rat Hippocampal Slices

Glutamate-mediated regulation of intracellular Ca²⁺ levels was examined in different populations of CA1 interneurons, using confocal microscopy and the Ca²⁺ indicator fluo 3-AM in rat hippocampal slices. Interneurons in basal [stratum oriendalveus (OA)] and apical [strata radiatum and lacunosum-moleculare (R/LM)] dendritic layers responded heterogeneously to glutamate. In control medium, OA interneurons responded mostly with oscillatory Ca²⁺ responses, which consisted of a large Ca²⁺ transient and successive smaller elevations. R/LM interneurons responded mostly with biphasic responses, characterized by an initial large transient and a secondary prolonged elevation. Other interneurons in both R/LM and OA responded with transient elevations in Ca²⁺ levels. lonotropic glutamate receptor antagonists (±)2-amino-5-phosphonopentanoic acid and 6-cyano-7-nitro-quinoxaline-2,3-dione reduced peak Ca²⁺ responses in OA and R/LM cells, and blocked biphasic responses in R/LM interneurons. The metabotropic glutamate receptor antagonist (RS)-α-methyl-4-carboxyphenylglycine reduced peak Ca²⁺ responses only in OA interneurons, and prevented oscillatory responses. In low Ca²⁺ medium, peak responses were reduced in R/LM but not in OA interneurons, and oscillatory responses were absent. Combination of ionotropic and metabotropic receptor antagonists blocked all glutamate-evoked Ca²⁺ responses. Activation of different types of glutamate receptors may thus produce heterogeneous Ca²⁺ signals in subpopulations of CA1 interneurons. lonotropic receptors may generate biphasic responses in interneurons in apical dendritic layers, whereas combined activation of metabotropic and ionotropic receptors may trigger oscillatory responses in interneurons of basal dendritic layers. These heterogeneous Ca²⁺ responses indicate that glutamate-mediated Ca²⁺ processes and second messenger systems differ in subpopulations of hippocampal interneurons and suggest possible postsynaptic functional specialization of interneurons.     

Paracoccidioidomycosis among the indian group Suruí of Rondonia, Amazonia, Brazil. A case report]

The Authors report a case of paracoccidioidomycosis in a Surui Indian patient from the state of Rondonia, Brazilian Amazon. The subject is an adult male, having been diagnosed on the basis of mycologic, serologic, and radiographic exams. The prescribed therapy was sulphamethoxazole 800 mg associated with trimethoprim 160 mg every 12 hours. A reevaluation of the patient conducted six months after the beginning of chemotherapy indicated overall improvement of his physical condition and of the radiologic picture, negativeness of the test of precipitin in capillary tube, and positiveness of the skin test with paracoccidioidin. The Authors also reviewed the cases of paracoccidioidomycosis described in the region, especially among the Surui Indian population.     

G→A mutations in p53 and Ha-ras genes in esophageal papillomas induced by N-nitrosomethylbenzylamine in two strains of rats

In human esophageal cancers, no ras gene mutations but a relatively high prevalence of p53 gene mutations have been reported. We found a high prevalence of point mutations in Ha-ras and p53 genes in N-nitrosomethylbenzylamine (NMBA)-induced esophageal tumors in two strains of rats (BD VI and F344). Our analysis showed the point mutation GGA←GAA (expected from the known mechanisms of action of NMBA) at Ha-ras codon 12 in 22 of 46 (48%) and 22 of 38 (58%) papillomas from BD VI and F344 rats, respectively. There was no significant difference in the prevalence of ras mutations in tumors induced by high doses (5.0 mg/kg) and low doses (2.5 mg/kg) of NMBA. Eleven papillomas from each strain were analyzed for p53 mutations. The prevalent mutations found were G←A and C←T transitions. The frequency of p53 mutation was 36% (four of 11) for each strain. No apparent hot-spot codon or exon was found in the p53 gene, and two papillomas contained double mutations in this gene. The high prevalence of G←A mutations in the rat Ha-ras gene contrasts with that in the human gene, in which no ras mutations have been found in primary tumors, and suggests either that the biology of esophageal carcinogenesis differs in humans and rats or that nitrosamines are not the major etiological risk factor for human esophageal cancers. © 1994 Wiley-Liss, Inc.     

Lithium transport in the mouse brain

Using the stable isotopes of lithium 6Li and 7Li, and the nuclear reaction 6Li(n,alpha)3H for detection, we have studied the isotopic exchange of lithium in various areas of the mouse brain and in the mouse plasma, under conditions of constant concentration of total lithium. The neutron irradiations were performed using 'cold' neutrons, at the European Institute Von Laue-Langevin. The nuclear reaction track densities were determined using an automatic image analyser. In the plasma, the isotopic ratios, 6Li/7Li, were measured using 'Secondary Ion Mass Spectrometry'. The concentration of total lithium in the plasma was kept close to 0.28 mM. The brain concentration of total lithium (referred to the tissue water content) ranged from more than 2 mM in the thalamus to less than 0.65 mM in the white matter of the cerebellum. The Nernst potential of lithium thus ranged from approx. -50 to approx. -20 mV, which means that lithium is probably not far from electrochemical equilibrium between brain cells and plasma. At any moment, the isotopic abundance of 6Li (ratio of 6Li to total lithium) in the different brain areas, were not significantly different from one another. The time-course of the isotopic abundance of 6Li in the brain was fitted by the composition of two exponential terms. The time-course of the isotopic abundance of 6Li in the plasma was also fitted by the composition of two exponential terms. These analytic curves (for the brain and for the plasma) were not significantly different from each other, at the precision of the measurements. This means that the isotopic equilibration of lithium between brain and plasma is almost instantaneous (i.e. accomplished in a few min at the most).     

Behavioral profile and dorsal hippocampal cells in carioca high-conditioned freezing rats

Selection for contextual fear conditioning is an important behavioral paradigm for studying the role of genetic variables and their interaction with the surrounding environment in the etiology and development of anxiety disorders. Recently, a new line of animals selectively bred for high levels of freezing in response to contextual cues previously associated with footshock was developed from a Wistar population. The purpose of the present study was to evaluate the emotional and cognitive aspects of this new line of animals, which has been named Carioca High-Freezing (CHF). For the characterization of anxious behavior, CHF and control animals were tested in the elevated plus-maze (EPM) and the social interaction test. CHF animals were significantly more anxious than control rats in terms of both the number of entries into EPM open arms and the percentage of time spent in these arms. The time spent in social interaction behavior was also significantly decreased. No statistical differences were found in locomotor activity, as measured by both the number of entries into the closed arms of the EPM and the number of crossings into the social interaction test arena. No differences between CHF and control groups were found in the depression forced swimming test, suggesting that the anxiety trait selected in the CHF line did not interact with affective disorders traits such as those for depression. Cognitive aspects of the CHF rats were evaluated in the object recognition task. Results from this test indicated no difference between the two groups. The present study also encompassed histological analysis of the dorsal hippocampus from CHF and control animals. Results revealed an absence of qualitative and quantitative differences between these two groups of animals in cells located in the dentate gyrus, CA1, and CA3 areas. Therefore, future studies are required to further investigate the possible neural mechanisms involved in the origin and development of the anxious phenotype observed in this model.     

Validation of a rhinitis symptom questionnaire (ISAAC core questions) in a population of Swiss school children visiting the school health services

The primary aim of the study was to assess the validity of the ISAAC core questions on rhinitis in a population of Swiss school children by comparing them to skin prick test results. Second, the positive predictive value in detecting atopy among children with rhinitis symptoms was determined. Third, agreement between parental reports of hay fever and rhinitis symptoms was evaluated, since earlier Swiss prevalence surveys had exclusively relied on reported hay fever. Material and methods: Two thousand nine hundred and fifty-four (81. 2%) parents of 7, 10 and 14-year old children filled in an exhaustive questionnaire which included the ISAAC core questions on rhinitis. Two thousand one hundred and twenty children also underwent skin prick testing against six common aeroallergens (grass mixture, birch, mugwort, D. pteronyssinus, cat and dog dander). The analysis is restricted to children with both questionnaire data and skin prick test results. Results: Sensitization to any allergen was most strongly associated with reported hay fever (OR = 5. 7, 95% CI 4. 4—7. 4), nose problems accompanied by itchy-watery eyes (OR = 4. 4, 95% CI: 3. 3—5. 7), symptoms occurring only during pollen season (March through September) (OR = 4. 9, 95% CI: 3. 6–6. 5) and a combination of these latter two symptoms (OR = 5. 8, 95% CI: 4. 1—8. 1). The association was stronger for a sensitization to outdoor allergens than for indoor allergens. The specificity of the various questions was high, ranging from 77. 5% to 97. 6%, but the sensitivity was low (2. 6% to 42. 7%). The positive predictive value for atopy among children with symptoms was 63% for sneezing accompanied by itchy-watery eyes, 67% for symptoms occurring only during the pollen season and 70% for reported hay fever. However, agreement between reptirted rhinitis symptoms and hay fever was only moderate. About one third of the children with symptoms indicative of seasonal rhinitis did not report the label “hay fever”. Conclusions: We conclude from our analyses that the ISAAC core questions on rhinitis are highly specific and therefore useful in excluding atopy. In addition they have a high positive predictive value in detecting atopy among children with symptoms, but they are not helpful for detecting atopy in a general population of children (low sensitivity). To monitor time trends in the prevalence of allergic rhinitis in Switzerland, questions on rhinitis symptoms as well as on the diagnostic label “hay fever” have to be included in a questionnaire because they contain complementary information since under-diagnosis of allergic rhinitis is common.     

Ten novel mutations in the HEXA gene in non-Jewish Tay -- Sachs patients

The heterogeneity of mutations causing Tay—Sachs diseasein non-Jewish populations requires efficient techniques allowingthe simultaneous screening for both known and novel mutations.ß-hexosaminidase mRNA isolated from cultured fibroblastsof 19 Tay-Sachs patients (7 with adult or late onset form ofthe disease and 12 with infantile Tay-Sachs disease) was amplifiedby cDNA—PCR in two overlapping segments spanning the entirecoding sequence. We used chemical mismatch cleavage (CMC), denaturinggradient gel electrophoresis (DGGE) and direct sequencing ofamplified fragments displaying a cleaved product or an alteredmelting behavior to screen the HEX A gene for mutations andto determine their distribution and frequency in the non-JewishTay—Sachs patients. These methods allowed us to identify31 out of 38 alleles studied (82%). In addition to 9 previouslydescribed mutations (the 4 bp insertion in exon 11, G to A transitionsat codons 170, 269, 482, 499 and 504, C to T transition at codon499 and 504 and a GT to AT transition at the donor site of intron9), we have identified 10 novel mutations. These include 1 donorsplice site defect in intron 6, 8 missense mutations at non-randomlydistributed conserved residues and a 2 bp deletion in exon 4.These results confirm the extreme molecular heterogeneity ofmutations causing Tay—Sachs disease in non-Jewish population.The strategy used should be profitable for identifying mutationsin large genes and for diagnostic purposes.     

Axonal Sprouting of CA1 Pyramidal Cells in Hyperexcitable Hippocampal Slices of Kainate-treated Rats

CA1 pyramidal cells become hyperexcitable following hippocampal kainate lesions. To examine if axonal sprouting contributes to this epileptiform activity, the local axonal arborization of CA1 pyramidal cells was examined after intracellular labelling with biocytin in hippocampal slices from control rats and in hyperexcitable slices obtained from rats treated with kainate (bilateral intracerebroventricular injections) 2-4 weeks previously. Biocytin-labelled cells with an axon that could be followed from the soma to the alveus were drawn and reconstructed with a camera lucida (15 cells from control slices and 14 cells from hyperexcitable slices). Local axonal arborizations were more extensive in cells of hyperexcitable slices. This increase in axon collaterals was generally seen in the alveus and in stratum oriens, but changes were more prominent in the latter. In stratum oriens, cells from hyperexcitable slices showed a significant increase in mean total axon length (1035 versus 373 μm in control), in mean number of branching points (6.50 versus 0.67 in control) and in mean number of segment orders per axon (3.07 versus 1.47 in control). Their first-order axon segments were similar in length to those of control cells (236 versus 338 pm in control), but with significantly more branching points (2.86 versus 0.53 in control). Their second-order axon segments were significantly longer (381 versus 63 μm in control) and also showed more branching points (2.71 versus 0.13 in control). Their third- and fourth-order axon segments were also longer and with more branching points. Under high-power light microscopic examination, biocytin-labelled axonal varicosities in cells of hyperexcitable slices were often seen in close apposition with their own dendrites, presumably making synaptic contact (five of nine cells examined). No such appositions were seen in any of the control cells (seven cells examined). These results indicate that, following kainate lesions, there is sprouting of local axon collaterals of CA1 pyramidal cells in stratum oriens and in the alveus. This local increase in axon collaterals may contribute to the epileptiform activity in the CA1 area by providing recurrent excitation via newly formed synaptic, and perhaps even autaptic, contacts with pyramidal cell dendrites.