Electron microscopy of the fate of exogenous ferritin in the feline visual cortex

Summary The ultrastructural changes occurring in the feline visual cortex 3 hours after the injection of 0.02 mls of ferritin in 1% trypan blue in artificial cerebrospinal fluid have been studied.Near the site of injection, disrupted cells contained free and membrane-bound ferritin. In less damaged areas, some signs of oedema were present in the cells, especially in astrocytes. Membrane-bound ferritin occurred occasionally in neurones and more frequently in astrocytes and oligodendrocytes. Considerable amounts of ferritin were also accumulated in phagocytic cells of unknown origin. In blood vessels, ferritin collected in the basement membrane and around collagen and, in membrane-bound form, in pale cells at the periphery of the vessels. Ferritin occurred in all parts of the intercellular space except in interglial junctions and tight junctions between vascular endothelial cells.This work was supported by a research grant from the National Health and Medical Research Council of Australia to Professor M. J. Blunt, of the School of Anatomy, University of New South Wales. The author wishes to thank Professor Blunt for his constant encouragement and support. The assistance of Mrs. Ruth Mather is gratefully acknowledged.     

The fine structure of the newborn rat adrenal cortex

Summary The structural changes of the zona juxtamedullaris of the rat adrenal cortex at birth, have been examined by the light and the electron microscope. In this zone clusters of medullary cells lying among the strands of cortical tissue were observed. In the inner portion of the zona juxtamedullaris two types of adrenocortical cells were found: light and very-dark cells. The latter are smaller than the light cells and are always in close connection with the medullary tissue. The ultrastructural features of the very-dark cells suggest that these elements are in degeneration. This finding supports the hypothesis that at birth there is a partial degeneration of the rat zona juxtamedullaris, i.e. the zone corresponding to the fetal zone of some mammalian species.It is proposed that in all mammalian species at birth there is a partial regression of the zona juxtamedullaris and that the regression of the fetal zone is only the quantitative increase of this phenomenon. This hypothesis is discussed in relation to numerous data demonstrating that there are enzymatic conditions in the rat during fetal life, which permit a discrete hypertrophy of the adrenal cortex.The author wishes to express his appreciation to Dr. A. Gambino and to Mr. G. Gottardo for technical assistance.     

Membrane Viscosity Correlates with α1-Adrenergic Signal Transduction of the Aged Rat Cerebral Cortex

We investigated, using adult (2-month-old) and senescent (12- and 24-month-old) rats, the effects of aging on the relationship between the alpha 1-adrenergic coupling system and the membrane viscosity of the cerebral cortex. There was no age-related difference in the KD values of [3H]prazosin binding on the membranes. The Bmax values of [3H]prazosin binding were reduced with advanced age. Norepinephrine-induced formation of 3H-labeled inositol phosphates (3H-IPs) in the slices increased with advanced age. The EC50 values for norepinephrine to stimulate the formation of 3H-IPs at advanced age were lower than that at adult age. The cholesterol content in membranes increased with advanced age. No changes in the phospholipid content in membranes were observed with advanced age. Concomitantly, an increase of the molar ratio of cholesterol to phospholipids was observed with advanced age. The membrane viscosity as measured by 1,6-diphenyl-1,3,5-hexatriene increased with advanced age. These results indicate that the altered cholesterol content and/or viscosity in cortical membranes of the aged rat may account for the loss of alpha 1-adrenergic receptor density and/or compensatory changes in the receptor-phospholipase C coupling system.     

Modulation of Rat Brain Cortical α-Adrenoceptors by Treatment with Hydrocortisone for 10 Days

The role of glucocorticoids in the modulation of central alpha 2-receptor mechanisms was investigated by in vitro receptor binding studies. [3H]Clonidine and [3H]idazoxan were used as radioligands. The alpha 2-receptor subtypes and guanine nucleotide sensitivity were studied in homologue and heterologue displacement experiments following hydrocortisone treatment (25 mg/kg s.c.) for 10 days. High and low agonist affinity states of the alpha 2-receptor could be identified in 3H-antagonist-agonist and 3H-agonist-antagonist displacement experiments, which may correspond to different regulatory protein-nucleotide associated forms of the receptor. In the presence of 10 microM GTP, the high-affinity binding was depressed. Following hydrocortisone treatment, there was no detectable change either in the affinity or the binding site concentration of clonidine in homologue displacement ("cold saturation") experiments. The affinity of idazoxan, however, was depressed. The effect of GTP was similar to the controls in this experimental arrangement. In contrast, in heterologue binding studies the high-affinity binding site was not demonstrable and the amount of low-affinity binding increased following the hydrocortisone treatment. The high-affinity site reappeared in the presence of GTP. The change in GTP sensitivity suggests that the nucleotide regulatory system may be involved in the action of adrenal steroids on central alpha 2-receptoral mechanisms.     

Comparison of [125I]Iodolysergic Acid Diethylamide Binding in Human Frontal Cortex and Platelet Tissue

The human platelet contains a functional 5-hydroxytryptamine (5-HT) receptor that appears to resemble the 5-HT2 subtype. In this study, we have used the iodinated derivative [125I]iodolysergic acid diethylamide ([125I]iodoLSD) in an attempt to label 5-HT receptors in human platelet and frontal cortex membranes under identical assay conditions to compare the sites labelled in these two tissues. In human frontal cortex, [125I]iodoLSD labelled a single high-affinity site (KD = 0.35 +/- 0.02 nM). Displacement of specific [125I]iodoLSD binding indicated a typical 5-HT2 receptor inhibition profile, which demonstrated a significant linear correlation (r = 0.97, p less than 0.001, n = 17) with that observed using [3H]ketanserin. However, [125I]iodoLSD (Bmax = 136 +/- 7 fmol/mg of protein) labelled significantly fewer sites than [3H]ketanserin (Bmax = 258 +/- 19 fmol/mg of protein) (p less than 0.001, n = 6). In human platelet membranes, [125I]iodoLSD labelled a single site with affinity (KD = 0.37 +/- 0.03 nM) similar to that in frontal cortex. The inhibition profile in the platelet showed significant correlation with that in frontal cortex (r = 0.96, p less than 0.001, n = 16). We conclude that the site labelled by [125I]iodoLSD in human platelet membranes is biochemically similar to that in frontal cortex and most closely resembles the 5-HT2 receptor subtype, although the discrepancy in binding capacities of [125I]iodoLSD and [3H]ketanserin raises a question about the absolute nature of this receptor.     

Age-dependent modifications of mitochondrial proteins in cerebral cortex and striatum of rat brain

The protein composition of free mitochondria purified from cerebral cortex and striatum during aging was analyzed by gel electrophoresis. Mitochondria were isolated from cerebral cortex and striatum of 4-, 12-, and 24-month-old rat brain. The percent amount of mitochondrial proteins after gel-electrophoretic separation was determined densitometrically. A significant decrease in the amount of two polypeptides (with molecular weights of 20 and 16 kDa, respectively) in both brain regions during aging was found. The decrease was higher in the striatum indicating a greater vulnerability of this brain area to the aging process. The age-dependent modifications of mitochondrial proteins observed may play an important role in several mitochondrial functions, such as energy transduction and transport processes as well as in structural changes occurring with age, causing altered membrane permeability and fluidity.     

Developmental changes in high-affinity uptake of GABA by cultured neurons

High-affinity uptake of [³H]-aminobutyric acid (GABA) was studied in cultures of neonatal rat cortical neurons grown on pre-formed monolayers of non-neuronal (glial) cells. Both the maximum rate (V _max) and, to a smaller extent, theK _m of [³H]GABA uptake increased with time. In addition, in parallel with these changes, 2,4-diaminobutyric acid and cis-3-aminocyclohexane-1-carboxylic acid (ACHC), compounds which are considered typical substrate/inhibitors of GABA uptake in neurons, became progressively stronger inhibitors of [³H]GABA uptake. Consequently, the present results may mean that the studies using uptake, of [³H]GABA, [³H]ACHC, or [³H]DABA as a specific marker for GABAergic neurons differentiating during the ontogenetic development of the central nervous system may have to be interpreted with caution.     

大鼠海马—小脑皮层—小脑深部核团投射的电生理学和HRP研究

本文用电生理学和HRP示踪法,研究了大鼠海马-小脑皮层投射的空间分布,小脑皮层的海马投射区与其深部核团间的纤维联系。 电生理学的实验结果表明,刺激背侧海马CA_1/CA_3区,均可使小脑皮层第Ⅵ小叶的浦肯野细胞产生顺行多突触的诱发简单锋电位和复杂锋电位反应。提示背侧海马CA_1/CA_3区与小脑皮层之间有经苔状纤维和攀缘纤维的多突触投射。实验证明,大鼠的这一投射的终止区域,集中在小脑皮层第Ⅵ小叶中线外侧0.8—1.4mm的范围内;并且来自CA_1区的投射以对侧性为主,CA_3区的投射以同侧性为主。HRP示踪的实验表明,背侧海马CA_1/CA_3区在小脑皮层第Ⅵ小叶的投射区是小脑纵区组构的间位区,该区皮层与间位核之间存在着交互投射关系。     

隐神经C类纤维传入诱发小脑皮层电反应

当弱刺激只引起隐神经A类纤维传入时,小脑皮层出现A-CEP,由潜伏期为11.8±3.5ms的早成分和312.1±17.5ms的晚成分组成;当强刺激同时引起A类和C类纤维传入时,出现AC-CEP类似A-CEP;用极化电流选择性阻断A类纤维传导后,只让C类纤维传入时,出现潜伏期为134.2±18.4ms的C-CEP。在Ⅵ小叶蚓部原裂附近C-CEP以正波为主,幅值最大,并在深层位相倒转。C-CEP的潜伏期较长,频率响应较低,幅值较小,随C类纤维传入量而变化,且对镇痛剂较敏感。结果表明C-CEP是由单纯C类纤维传入引起的,在小脑皮层内产生,是小脑皮层对慢痛信息传入的反应。提示C类纤维传入可以到达小脑皮层,引起诱发电位。当A和C类纤维同时传入时,C-CEP不出现,可能是被A类纤维传入所抑制。     

Regulation of the spatial order of cortical microtubules in developing guard cells ofAllium

The organization of microtubules (MTs) in the cortex of cells at interphase is an important element in morphogenesis. Mechanisms controlling the initiation of MTs and their spatial ordering, however, are largely unknown. Our recent study concerning the generation of a radial array of MTs in stomatal guard cells inAllium showed that the MTs initiate in a cortical MT-organizing zone adjacent to the ventral wall separating the two young guard cells (Marc, Mineyuki and Palevitz, 1989, Planta179, 516, 530). In an attempt to detect MT-ordering mechanisms separate from the sites of MT initiation, we now employ various drugs to manipulate the geometry and integrity of the ventral wall and thereby also the associated MT-organizing zone. In the presence of cytochalasin D the ventral wall is tilted away from its normal mid-longitudinal anticlinal alignment, while treatments with the herbicide chloroisopropyl-N-phenylcarbamate (CIPC) induce the formation of a branched ventral wall. Nonetheless, in either case the MTs still form a radial array, although this is asymmetric as it is centered in accordance with the misaligned or branched ventral wall. Since the MTs maintain their original course undisturbed as they extend beyond the abnormal ventral wall, there is no evidence for the presence of an inherent MT-ordering mechanism at locations remote from MT-initiation sites. Following treatments with caffeine, which abolishes the formation of the ventral wall, the MTs revert to a transversely oriented cylindrical array as in normal epidermal cells. Thus the presence of the ventral wall, and presumably also the associated MT-organizing zone, is essential for the establishment of the radial array. The MT-organizing zone is therefore involved not only in the initiation of MTs, but also in determining their spatial order throughout the cell cortex. We thank Drs. Richard J. Cyr and Yoshi Mineyuki for providing valueable suggestions during the course of this work, and Ms. Elizabeth Bruce printing some of the figures. This research was supported by Funds from the National Science Foundation grants DCB-8703292 to B.A.P. and DCB-8803286 to B.A.P. and J.M.