The laminar organization of the motor cortex in monodactylous mammals: a comparative assessment based on horse, chimpanzee, and macaque.

The architecture of the neocortex classically consists of six layers, based on cytological criteria and on the layout of intra/interlaminar connections. Yet, the comparison of cortical cytoarchitectonic features across different species proves overwhelmingly difficult, due to the lack of a reliable model to analyze the connection patterns of neuronal ensembles forming the different layers. We first defined a set of suitable morphometric cell features, obtained in digitized Nissl-stained sections of the motor cortex of the horse, chimpanzee, and crab-eating macaque. We then modeled them using a quite general non-parametric data representation model, showing that the assessment of neuronal cell complexity (i.e., how a given cell differs from its neighbors) can be performed using a suitable measure of statistical dispersion such as the mean absolute deviation-mean absolute deviation (MAD). Along with the non-parametric combination and permutation methodology, application of MAD allowed not only to estimate, but also to compare and rank the motor cortical complexity across different species. As to the instances presented in this paper, we show that the pyramidal layers of the motor cortex of the horse are far more irregular than those of primates. This feature could be related to the different organizations of the motor system in monodactylous mammals.

Gene expression profiles of estrogen receptors α and ß in the fetal bovine hypothalamus and immunohistochemical characterization during development.

Steroid hormones intervene in the structural and functional regulation of neuronal processes during development and thus determine brain differentiation. The effects of estrogens are mediated by two transcription factors, namely estrogen receptor α (ER-α) and estrogen receptor ß (ER-ß), that regulate the expression of target genes through their binding to specific DNA target sequences. We describe the mRNA expression of ER-α and ER-ß in the hypothalamus of developing male and female bovines as revealed by quantitative real-time polymerase chain reaction, and the distribution of the two ERs in hypothalamic sections of all fetal stages as shown by immunohistochemistry. The expression profiles of the mRNAs of both ERs are mutually correlated throughout the gestation period, and their levels increase significantly in the last stages of gestation. No sexual differences in the mRNA expression of either ER-α or ER-ß have been found in our fetal specimens. The use of specific antisera against ER-α and ER-ß has allowed us to characterize and confirm the distribution of these receptors in the hypothalami of all fetal stages considered. Our results offer detailed information concerning the distribution of ER-α and ER-ß in the developing bovine hypothalamus and provide additional insights into the processes involved in the hypothalamic development of a mammal with a long gestation and a highly gyrencephalic brain.

Expression profile of the pore-forming subunits α1A and α1D in the foetal bovine hypothalamus: a mammal with a long gestation.

This study describes the expression of the voltage operated calcium channels (VOCCs) subunits α1A (typical of the P/Q family) and α1D (of the L family) in the bovine hypothalamus. The expression of both P/Q and L families has been characterized in the brain of adult mammals. However, their distribution and expression during foetal neuronal differentiation have not yet been determined. The expression profile of the α1A and α1D pore-forming subunits was investigated during four embryonic stages in bovine foetuses. Our data suggest that the expressions of α1A and α1D are correlated during development, with an increase only in males that peaks on the last period of gestation. Bovine male hypothalami showed significantly higher α1A and α1D expression values in comparison to female ones during the whole developmental period. In the females, the expression profiles of both genes were constant during all the developmental time. Immunohistochemical studies confirmed the presence of the α1A and α1D protein subunits in foetal hypothalamic neurones starting from the third foetal stage. Our data provide new information on the hypothalamic expression of α1A and α1D subunits during development in a mammal with a long gestation period and a large and convoluted brain.

Characterization of an established endothelial cell line from primary cultures of fetal sheep hypothalamus.

Immortalized cell lines from fetal brain are an experimental model for studying the in vitro molecular pathways regulating neural cell differentiation and the development of neural networks. The procedures are described to obtain an established cell line from the 90-day old fetal sheep hypothalamus. Viral oncogene LT-SV40 transformation was used to isolate a stable cell line (ENOS-01) that was characterized immunocytochemically. Immortalized cells can be classified as an endothelial cell line of hypothalamic microvasculature. Furthermore, mRNA expression and immunocytochemical of estrogen receptors α and ß were also evaluated. Since it is known that cerebral vessels are directly targeted by sex steroids, our established cell line represents an alternative system to study estradiol/receptor interactions during brain development. Our in vitro model can provide a tool to investigate the complex relationships among the cell types forming the blood-brain barrier, which is known to be involved in the pathogenesis of sheep transmissible neurological diseases.

Expression of aromatase P450(AROM) in the human fetal and early postnatal cerebral cortex.

Aromatase (P450(AROM)), the enzyme responsible for the conversion of testosterone (T) into 17-ß estradiol (E(2)), plays a crucial role in the sexual differentiation of specific hypothalamic nuclei. Moreover, recent findings indicate that local E(2) synthesis has an impact on other brain areas including hippocampus, temporal cortex and cerebellum, and may thus influence also cognitive functions. Numerous studies have described the expression and the distribution of P450(AROM) throughout ontogenesis and postnatal development of the central nervous system in several mammals, but data referring to humans are scarce. In the adult human brain, P450(AROM) has been detected in the hypothalamus, limbic areas, and in the basal forebrain, and described in glial cells of the cerebral cortex and hippocampus. In this study we report the expression, distribution and cellular localization of P450(AROM) in the human fetal and early postnatal cerebral cortex. In our series of fetal brains of the second trimester, P450(AROM) expression appeared at gestational week (GW) 17 and resulted limited to groups of cells localized close to the growing neuroepithelium in the ventricular and subventricular zones. At GWs 20-24, scattered P450(AROM) immunoreactive (-ir) neural cells were identified in the intermediate plate and subplate, and in the parietal cortical plate. In perinatal and early postnatal individuals the quantity of P450(AROM)-ir elements increased, and revealed the morphology typical of glial cells. Double labeling immunostaining with anti-GFAP and anti-P450(AROM) antisera, and subsequent confocal analysis, confirmed this observation. Our data show that the expression of P450(AROM) in the fetal cortex starts approx at the end of the fourth gestational month, but increases steadily only in the last trimester or in the early postnatal period. This temporal trend may suggest that P450(AROM) could act as a differentiation-promoting factor, based on timing of the steroid actions.

Expression and localization of aromatase P450AROM, estrogen receptor-α, and estrogen receptor-ß in the developing fetal bovine frontal cortex.

The enzyme aromatase (P450(AROM)) converts testosterone (T) into 17-ß estradiol (E(2)) and is crucial for the control of development of the central nervous system during ontogenesis. The effects of E(2) in various brain areas are mediated by the estrogen receptor alpha (ER-α) and the estrogen receptor beta (ER-ß). During fetal development, steroids are responsible for the sexual differentiation of the hypothalamus. Estrogens are also able to exert effects in other brain areas of the fetus including the frontal cortex, where they act through estrogen receptors (ERs) modulating cognitive function and affective behaviors. In this study we have determined the expression profiles of P450(AROM) and ERs in the fetal bovine frontal cortex by quantitative Real-Time PCR (qRT-PCR) throughout the prenatal development. The data show that the patterns of expression of both ERs are strongly correlated during pregnancy and increase in the last stage of gestation. On the contrary, the expression of P450(AROM) has no correlation with ERs expression and is not developmentally regulated. Moreover, we performed immunochemical studies showing that fetal neurons express P450(AROM) and the ERs. P450(AROM) is localized in the cytoplasm and only seldom present in the fine extensions of the cells; ER-α is detected predominantly in the soma whereas ER-ß is only present in the nucleus of a few cells. This study provides new data on the development of the frontal cortex in a long gestation mammal with a large convoluted brain.

Correlation of minimum coronary lumen diameter with left ventricular functional impairment induced by atrial pacing.

To understand whether quantitative measurement of minimal coronary luminal diameter is a better method than percent diameter narrowing for assessing the functional impairment of myocardial contractility produced by coronary artery stenoses, measurements were made from 37 stenotic segments in 27 patients with coronary artery disease and from corresponding segments in 10 subjects without coronary artery narrowing. An assessment of the reliability of the 2 types of measurements was made by correlating them with the physiologic parameters of both segmental wall motion and global ejection fraction response induced by atrial pacing. Digitally acquired coronary angiograms were used to facilitate quantitative analysis. Measurements by edge detection and videodensitometry correlated closely (r = 0.94). Percent diameter narrowing correlated moderately with the change in ejection fraction (r = -0.41) or with the change in segmental wall motion (r = -0.44). The measurement of minimal lumen diameter correlated with the change in global ejection fraction (r = 0.61) and did so even better with the change in segmental wall motion (r = 0.78, p less than 0.05). A minimal lumen diameter of less than or equal to 1.5 mm identified patients likely to have a functional impairment during atrial pacing as assessed by either global ejection fraction or segmental wall motion defects. We conclude that minimal coronary luminal diameter provides a better method than percent diameter narrowing calculations to measure the anatomic severity of coronary artery narrowing.

Videodensitometric determination of minimum coronary artery luminal diameter before and after angioplasty.

Quantitative measurements of coronary stenoses were made from digital coronary angiograms in 19 patients before and after percutaneous transluminal coronary angioplasty (PTCA). Two methods of measurement were compared. Mean stenosis before PTCA was 67 +/- 10% by the edge detection method and 67 +/- 12% by videodensitometry (difference not significant). After PTCA, the mean stenosis was 32 +/- 14% by edge detection and 30 +/- 13% by videodensitometry (difference not significant). In addition, a new method was developed to rapidly calculate the absolute minimum luminal area and diameter by videodensitometry. The minimum luminal diameter before PTCA was 1.0 +/- 0.5 mm and after PTCA increased to 2.4 +/- 0.5 mm (p less than 0.001). The validity of the videodensitometric method was analyzed in a series of Lucite phantom studies, which suggested that when there is an irregular angiographic appearance, the densitometric method may be more accurate than standard edge detection methods. Digital acquisition of coronary angiograms provides a means for rapid application of quantitative analysis during coronary interventional procedures.

Digital coronary roadmapping as an aid for performing coronary angioplasty.

In an attempt to improve visualization of the position of the guidewire and dilatation balloon during coronary angioplasty, a method was developed called digital coronary roadmapping. With this method a digitally acquired coronary angiogram is interlaced with the live fluoroscopic image of the guidewire and balloon catheter. The digital coronary angiogram is superimposed at the same magnification and radiologic projection as the live fluoroscopic image onto the video monitor above the catheterization table. The digital roadmap image thus provides immediate feedback to the angiographer to assist in directing the guidewire into the appropriate coronary artery branch and to help in placement of the balloon so that it straddles the site of stenosis.

Detection and quantitation of coronary artery stenoses from digital subtraction angiograms compared with 35-millimeter film cineangiograms.

To assess the ability to detect coronary artery narrowings from computer-acquired angiograms, a panel of 4 observers independently identified and measured focal coronary narrowings from digital subtraction angiograms and compared the results to those obtained from standard 35-mm cine film angiograms. Both cine and digital angiograms were obtained sequentially using selective intracoronary artery injection of standard amounts of iodinated contrast media. Digital images were obtained at 8 frames/s with a 512 X 512 X 8-bit pixel matrix. Modifications in the imaging chain for computer acquisition included a slower pulsed radiographic mode, a progressive scan camera, and initial storage of the images on an 80-megabyte digital hard disk. Postprocessing computer algorithms were used to enhance the unsubtracted digital images; these included single-frame, mask-mode subtraction, vessel boundary edge enhancement, and 4-fold pixel magnification. In 19 patient studies, 32 arteries were reduced more than 25% in diameter according to at least 1 of 4 observers on either the digital or cine film angiograms. There was no significant difference in the mean percent diameter narrowing for all the narrowings between the digital angiograms (53 +/- 31%) and the cineangiograms (52 +/- 31%). In addition, a 2-way analysis of variance yielded no significant difference between the amount of variability in the measurements between the cine film and the digital technique. This similar variability persisted when subsets of patients based on the degrees of stenosis were considered (e.g., only narrowings from 50 to 90% diameter reduction).(ABSTRACT TRUNCATED AT 250 WORDS)