Role of medical history in brain tumour development. Results from the international adult brain tumour study.

In an international population-based case-control study carried out in 8 centres in 6 countries, we investigated the role of specific medical conditions in the aetiology of brain tumours in adults. Recruited were 1,178 glioma and 331 meningioma cases and 2,493 age- and gender-matched population controls. Only medical conditions occurring at least 2 years before brain tumour diagnosis were considered. Relative risks (RRs) and 95% confidence intervals (CIs) were estimated using a conditional logistic regression model. Heterogeneity between centres was tested. No association between meningioma and previous medical conditions was observed. For glioma, there was an increased risk associated with epilepsy (RR = 6.55, 95% CI 3.40-12.63), but this was considerably weaker for epilepsy of more than 20 years duration. The risk remained elevated after adjustment for use of anti-epileptic drugs. There was a statistically significant inverse association between glioma and all allergic diseases combined (RR = 0.59, 95% CI 0.49-0.71); this was also observed for specific allergic conditions, namely, asthma and eczema. Subjects who reported a history of infectious diseases (e.g., colds, flu) showed a 30% reduction in risk (RR = 0.72, 95% CI 0.61-0.85). The decreased risks for glioma in subjects reporting a history of allergic conditions or infectious diseases may indicate an influence of immunological factors on the development of glioma. The association between glioma and epilepsy has to be interpreted cautiously and needs further investigation.

Sero-epidemiological analysis of the risk of virus infections for childhood leukaemia.

Virus infections have been thought to be involved in the development of childhood leukaemia. In order to address this issue we determined, in a case-control study, the prevalence of antibodies to viruses infecting blood or bone-marrow cells [Epstein-Barr virsus (EBV), human herpes virus type 6 (HHV-6), parvovirus B19] as well as to the human virus known for its tumour-suppressive properties, the adeno-associated virus type 2 (AAV-2), in the sera of 121 children with leukaemia in Germany, and in 197 control individuals, hospitalized for other reasons, and matched for age and gender to the cases. In addition, we developed a questionnaire to be answered by the children's parents, in order to gain information on previous infections of the children as well as to calculate for factors which may influence serological findings. Comparative determination of the prevalence of antibodies against AAV-2, B-19 or HHV-6 revealed no significant differences in cases and controls. However, antibodies to EBV were more frequently found in children with leukaemia younger than 6 years of age (age at the time of diagnosis of leukaemia) than in controls. Apparently, infection with AAV-2 has no protective effect in childhood leukaemia, in contrast to results observed for other malignancies. Similarly, and in accordance with results on leukaemia in adults, we found no indication of a protective effect of infection with the parvovirus B-19. The data suggest that EBV, which is known to be involved in various lymphomas, may play a role in the development of childhood leukaemia in young children.

Occupation, smoking and demographic factors, and renal cell carcinoma in Germany.

BACKGROUND: The role of occupational exposure in the aetiology of renal cell cancer is still not clear. In a population-based, case-control study we investigated occupational and smoking history as well as place of residence, marital and socioeconomic status. METHOD: In a case-control study in Germany, 277 adult cases with incident renal cell cancer and 286 controls frequency-matched to the cases for age and gender have been interviewed. The data were analysed by standard methods using unconditional logistic regression models, to estimate the relative risk (RR) and the corresponding 95% confidence interval (95% CI). RESULTS: Socioeconomic status was inversely associated (RR = 0.61, 95% CI: 0.3-1.2 for highest category) with the risk for renal cell cancer. Heavy smoking gave an increased, but not significant (about twofold) RR in men and women for ex-smokers and current smokers. Employment in metal-related industries (RR = 1.63, 95% CI: 1.1-2.5) was also identified as a risk factor. Additionally, we found an elevated risk associated with exposure to perchloroethylene and tetrachlorocarbonate (RR = 2.52, 95% CI: 1.2-5.2) but no time trend could be observed. No associations were found for other occupational exposures, such as working in the chemical industry, transportation or farming nor for exposure to pesticides. CONCLUSIONS: The results of our study lead to the suggestion that smoking, occupation and demographic factors probably play a minor role in the aetiology of renal cell cancer.

Occupational risk factors for brain tumors: results from a population-based case-control study in Germany.

In a population-based case-control study in the Rhein-Neckar-Odenwald area (containing 1.3 million inhabitants) of the Federal Republic of Germany (FRG), risk factors were assessed for brain tumor development in 226 cases with primary brain tumors (ICD-9 191, 192.1, 192.0) and 418 population controls, interviewed by a standardized questionnaire. The analysis of occupational risk factors and smoking is presented. No elevated risk was found for smoking. Similarly, no significant effects were found for most occupations. Five specific occupational groups were examined because of a priori determination that they were of interest. Some categories showed slightly elevated risks but in none was the elevation statistically significant. A significant increase in risk for brain tumor development was found associated with working in electrical occupations for women (relative risk [RR] = 5.2; 95 percent confidence interval [CI] 1.4-20.1) but not for men (RR = 0.9, 95 percent CI 0.3-2.3).

Quantitative autoradiographic localization of cholecystokinin receptors in rat and guinea pig brain using 125I-Bolton-Hunter-CCK8.

The autoradiographic localization of receptors for the brain-gut peptide cholecystokinin (CCK) has shown differences in receptor distribution between rat and guinea pig brain. However the full anatomical extent of the differences has not been determined quantitatively. In the present study, 125I-Bolton-Hunter-CCK8 (125I-BH-CCK8) was employed in a comparative quantitative autoradiographic analysis of the distribution of CCK receptors in these two species. The pharmacological profile of 125I-BH-CCK8 binding in guinea pig forebrain sections was comparable to those previously reported for rat and human. Statistically significant differences in receptor binding between rat and guinea pig occurred in olfactory bulb, caudate-putamen, amygdala, several cortical areas, ventromedial hypothalamus, cerebellum, and a number of midbrain and brainstem nuclei. The results of this study confirm the presence of extensive species-specific variation in the distribution of CCK receptors, suggesting possible differences in the physiological roles of this peptide in different mammalian species.

Somatostatin alters beta-adrenergic receptor-effector coupling in cultured rat astrocytes.

The neuropeptide somatostatin potentiates beta-adrenergic receptor-mediated cAMP formation in astrocytes derived from neonatal rat cortex but does not affect cAMP levels by itself. beta-Adrenergic receptors in these cells can be specifically labeled with the high affinity antagonist [125I] cyanopindolol ([125I]CYP). In addition, astrocytes display both high and low affinity binding sites for the agonist isoproterenol, which are thought to represent receptors which are coupled or uncoupled, respectively, to the guanine nucleotide regulatory protein. We find that somatostatin does not modify beta-receptor density, nor receptor affinity for either the antagonist ([125I]CYP) or for the agonist isoproterenol. In the presence of the guanine nucleotide analogue, Gpp(NH)p, only low affinity (uncoupled) displacement of [125I]CYP binding by isoproterenol is observed. However, somatostatin (1 microM), when added to the cells together with Gpp(NH)p, prevents the nucleotide-induced loss of the high affinity (coupled) component of agonist displacement. This result suggests that somatostatin increases noradrenaline-induced cAMP production by enhancing coupling between the beta-receptor and the stimulatory guanine nucleotide regulatory protein.

Differential localization of type I and type II benzodiazepine binding sites in substantia nigra.

A number of studies have suggested the existence of multiple benzodiazepine binding sites in the brain. We have recently reported the physical separation of two apparent benzodiazepine binding site subtypes, the pharmacological properties, and distribution in tissue sections of which correspond to the putative type I and type II sites. Benzodiazepine and gamma-aminobutyric acid (GABA) receptors have been shown to interact, and lesions of the GABAergic striatonigral pathway, which lead to GABA supersensitivity, both increase the numbers of GABA binding sites and enhance GABA-stimulated benzodiazepine binding. We demonstrate here that degeneration of striatonigral fibres increases the density of putative type I benzodiazepine binding sites in the substantia nigra and decreases the density of the putative type II sites. This suggests that type I sites that increase after denervation are postsynaptic, whereas the type II sites reduced by the lesion may be localized to axons or terminals of the striatonigral pathways.

Normalization of spiroperidol binding in the denervated rat striatum by homologous grafts of substantia nigra.

Transplantation of embryonic substantia nigra into the adult rat brain decreases the motor asymmetry that is produced by dopamine receptor supersensitivity after a unilateral lesion of the substantia nigra. The authors report that this effect of transplantation is specific to grafts of substantia nigra. They also report that, in conjunction with the decrease in motor asymmetry, these grafts cause postsynaptic dopaminergic binding sites to return to normal density as measured by tritiated spiroperidol autoradiography. Thus, in animals with brain lesions, grafts of substantia nigra produce a long-term alteration in the functional status of host brain cell receptors that is associated with a reduction in the behavioral deficit.

Benzodiazepine receptors: localization in rat amygdala.

Behavioral, electroencephalographic, and evoked potential studies, as well as experiments utilizing the direct intracerebral injection of benzodiazepines (BZs), have implicated the amygdala, particularly its basolateral nuclear division, as an important locus of the therapeutic actions of these drugs. BZ receptor localization within the amygdala could further define which amygdaloid nuclei are important for BZ action and suggest neuroanatomical pathways especially sensitive to these drugs. Using a quantitative light microscopic autoradiographic technique, this study has localized type 1 and type 2 BZ receptors within the amygdaloid nuclear complex of the rat. BZ receptors were labeled with [3H]flunitrazepam, and type 1 BZ receptor binding was preferentially displaced by CL218,872 in 8-micron coronal sections of rat brain. Autoradiograms were generated by the apposition of emulsion-coated coverslips and quantified by standardized procedures. Ten amygdaloid nuclei were examined at nine levels over the rostrocaudal extent of the amygdala. Total BZ receptor density was highest in the basolateral nuclear complex, and the majority of these receptors were of the type 2 subclass. Type 1 receptors were concentrated in the anterior aspects of the amygdala, particularly the anterior cortical nucleus. These results are consistent with an important role for the basolateral amygdala in anxiogenesis and seizure induction. Furthermore, a larger circuit which includes the amygdala, ventromedial hypothalamus, mammillary body, anterior thalamus, and frontal cortex could be involved in the anxiolytic actions of the BZ drugs.

Multiple benzodiazepine receptors: autoradiographic localization in normal human amygdala.

Because previous studies have emphasized the importance of the amygdala in the therapeutic actions of benzodiazepines and described differences in benzodiazepine receptor distribution between human and rat, this study examined the distribution of multiple benzodiazepine receptors in normal human amygdala by light microscopic autoradiography. Benzodiazepine receptors were labeled with [3H]flunitrazepam at 5 representative rostro-caudal levels. Type 1 and Type 2 receptors were differentiated by the addition of 200 nM CL218,872 to serial sections and subsequent analysis based on the differential occupancy at Type 1 and Type 2 receptors by this drug. Results demonstrated that like the rat amygdala, human amygdala contains a higher density of benzodiazepine receptors in the basolateral nuclear complex compared to the corticomedial complex, and more Type 2 than Type 1 receptors overall. However, while rat amygdala is enriched rostrally in Type 1 receptors, this subclass in humans is elevated caudally. Pathways including the amygdala and limbic and cortical targets of its efferents may be preferential loci of benzodiazepine anxiolytic activity.

Autoradiographic differentiation of multiple benzodiazepine receptors by detergent solubilization and pharmacologic specificity.

We have used light microscopic autoradiography to localize differentially Type 1 and 2 benzodiazepine receptors using pharmacologic and detergent treatment techniques. The triazolopyridazine CL218872 has preferential affinity for Type 1 receptors from which it displaces [3H]flunitrazepam selectivity. In biochemical experiments [3H]flunitrazepam binding sites with pharmacologic properties of Type 2 receptors are preferentially solubilized from brain membranes by detergents. We have now treated rat brain slices either with CL218872 or 2% sodium cholate and evaluated the autoradiographic distribution of [3H]flunitrazepam-labeled receptors. Whether calculated from the drug or detergent treated preparations, the relative densities of apparent Type 1 and 2 receptors are the same. Thus differential sensitivity to determine solubilization distinguishes the same two populations of receptors as Type 1 and 2 specific drugs.

Benzodiazepine receptors: preferential stimulation of Type 1 receptors by pentobarbital.

The binding of [3H]flunitrazepam (FLU) to Type 1 and Type 2 benzodiazepine receptors in rat cerebellum and cerebral cortex was differentiated by the addition of 200 nM CL218,872 which preferentially displaces [3H]FLU from Type 1 receptors. Type 1 but not Type 2 receptor binding was significantly stimulated by 1 mM sodium pentobarbital.

Quantitative receptor autoradiography using [3H]ultrofilm: application to multiple benzodiazepine receptors.

Radiohistochemical methods utilized to label drug and neurotransmitter receptors in vitro at the light microscopic level are quantitative. In this report, the response of tritium-sensitive sheet film ([3H]Ultrofilm) was characterized under conditions used for the light microscopic localization of neurotransmitter receptors by in vitro autoradiographic techniques. Radioactive standards containing varying concentrations of tritium were prepared from brain tissue and exposed to [3H]Ultrofilm for varying lengths of time, and the response of the film was measured by microdensitometry. A 1n optical density versus 1n radioactivity plot provided a useful linear relationship. Relationships established between radioactivity concentrations, exposure times and optical density were utilized to establish appropriate exposure conditions for several [3H]ligands in brain sections. The accuracy and pharmacological relevance of these methods were tested by studying the regional distribution of multiple benzodiazepine (BZ) receptors, and by analyzing the inhibitory potency of the triazolopyridazine, CL218,872, and methyl-beta-carbolinecarboxylate, two agents which discriminate between type 1 and 2 BZ receptors. The results obtained compared favorably with results previously obtained in tissue homogenates by biochemical methods. Overall, these results have practical implications. For example, quantitative autoradiography can be used to generate accurate kinetic and pharmacological displacement curves for small tissue areas. Also, since the film response curve is not linear with exposure (radioactivity X time), autoradiographs of the same sections can have relatively different density patterns depending on the time of exposure.

Binding of a radiolabeled triazolopyridazine to a subtype of benzodiazepine receptor in the rat cerebellum.

The triazolopyridazine (TPZ) drugs, typified by CL218,872 (CL), have a relatively higher affinity for a subpopulation of benzodiazepine (BZ) receptors. The binding of radiolabeled CL to membranes from rat cerebellum, a region enriched in the TPZ-preferring ("Type 1") BZ receptor, was characterized and compared with that of [3H]flunitrazepam ([3H]FLU) in the same preparation. [3H]CL had clonazepam displaceable binding which was saturable. The Kd was approximately 21 nM and the Bmax was approximately 600 fmol/mg of protein. [3H]CL binding was similar to that for [3H]FLU in that exogenous gamma-aminobutyric acid (GABA) enhanced the binding; however, [3H]CL binding differed from that for [3H]FLU in that anions, cartazolate and pentobarbital did not enhance [3H]CL binding. These data suggest that [3H]CL binds to the Type 1 BZ receptor in a manner different from that of a BZ drug such as FLU. Inasmuch as GABA enhances [3H]CL binding, but anions, cartazolate and pentobarbital do not, [3H]CL may bind to the Type 1 BZ receptor in such a way that it interacts with the GABA site, but perhaps not directly with the ionophore or the postulated pyrazolopyridine-barbiturate site. Thus, TPZ drugs may affect the GABA receptor complex in a different or perhaps less extensive way than the BZs. This, in addition to the regional localization of the Type 1 receptor, may be an important part of the mechanism of action of the TPZs.

Receptor autoradiography with tritium-sensitive film: potential for computerized densitometry.

Benzodiazepine receptors mounted tissue sections of brain were localized by autoradiography using a tritium-sensitive sheet film (3H-Ultrofilm, LKB). The use of the film had many advantages compared to a previous technique utilizing emulsion-coated coverslips. A major advantage is the adaptability of the film to microdensitometric techniques for measurement of receptors. The densitometry can be carried out by computerized, color-coded image analysis.

Benzodiazepine receptors are coupled to a subpopulation of gamma-aminobutyric acid (GABA) receptors: evidence from a quantitative autoradiographic study.

The colocalization and interaction of gamma-aminobutyric acid (GABA) and benzodiazepine (BZ) receptors in the rat brain were characterized using standardized, quantitative, light microscopic autoradiographic methods. In serial sections, striking differences were observed in the distribution of high affinity GABA and BZ receptors in areas such as the cerebral cortex, globus pallidus, thalamus, hypothalamus and cerebellar cortex. However, in a semiquantitative visual examination of more than 200 brain regions, added exogenous GABA increased BZ binding in all regions. In a quantitative analysis of 19 regions, exogenous GABA uniformly stimulated [3H]flunitrazepam binding, the effect being proportional to the regional density of BZ receptors. No relationship was seen between the magnitude of the stimulation and the distribution of high affinity GABA receptors. In the mounted tissue sections, BZ binding appeared influenced by endogenous GABA since it was reduced by preincubation or by the addition of bicuculline. Taken together, these data suggest that most or all BZ receptors can be influenced by GABA and are coupled to a type of GABA receptor. However, the BZ-linked GABA receptor could represent either a subpopulation of GABA binding sites or a distinct receptor not labeled under the conditions used in these and other experiments.

[3H]Spiperone binding sites in brain: autoradiographic localization of multiple receptors.

[3H]Spiperone ([3H]SP) binding sites were localized by light microscopic autoradiography, after in vitro labeling. The kinetic and pharmacological characteristics of these binding sites were studied in slide-mounted sections of rat forebrain, and optimal labeling conditions were defined. Autoradiograms were obtained by apposing emulsion-coated coverslips to labeled sections. Differential drug sensitivity allowed the selective displacement of [3H]SP from dopamine receptors by ADTN, from serotonin receptors by cinanserin, from both by haloperidol and from unique spiperone sites by unlabeled spiperone. The various sites presented a differential anatomical localization. For example, only dopaminergic sites were found in the glomerular layer of the olfactory bulb; only serotonergic sites were found in lamina IV of the neocortex, and a high concentration of unique spiperone sites was found in parts of the hippocampus.