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The cascade of pathobiochemical reactions caused by cerebral ischemia is complex and not completely understood. One important factor seems to be the disturbed calcium homeostasis. Measurement of cytosolic-free Ca2+ concentration [Ca2+]i is possible with the Ca(2+)-sensitive dye fura-2. Using this technique it can be investigated if, and how, neuroprotective drugs influence the cellular calcium homeostasis. In hippocampal neurons K+ or glutamate, added to the buffer solution, evoked elevations of [Ca2+]i in a concentration-dependent manner. We demonstrated that the calcium antagonist nimodipine or the non-competitive NMDA receptor antagonist dizocilpine added to the buffer solution could inhibit concentration-dependently the elevation of [Ca2+]i. The measurement of [Ca2+]i can be used for studies on the mechanism of action as well as for screening purposes of neuroprotective drugs.

Regarding the hippocampal formation and especially the external two thirds of it's dentate molecular layer a lot of possible morphological changes after long-term potentiation (LTP) have been described in literature. The present morphometric-stereological study of vesicles in axo-spino-dendritic synapses of the inner third of the molecular layer was done under the aspect of heterosynaptic influences following LTP. Because of the hierarchical link of the three analytic levels (test-group, animal, synapse), for statistical interpretation we used the analysis of variance with two-way hierarchical classification. Between the 3 groups (passive control, active control, LTP-group) we found no significant differences. Because of the great differences between the vesicles even within a single synapse we subsequently investigated the middle third of the molecular layer, i.e. the terminal area of the stimulated perforant path. No differences between the three groups we found here either. There was no confirmation for the expected greater homogenization of the synapses based on the uniform input. As a result of this study pure morphological studies without selective staining of specific population of synapses are considered inadvisable. Only with the help of selective staining in the area of the synapses possible differences between the groups may be found.

In this paper we have investigated in 20 Wistar rats the projection of the locus coeruleus (LC) and of the two mesencephalic raphe nuclei Ncl. raphe centralis superior (NCS) and Ncl. raphe dorsalis (NRD) to the hippocampus by using the HRP-technique. 1. The noradrenergic projection of the LC courses mainly ipsilaterally. On the contralateral side their are in the LC only a few labelled cells (about 10% of the labelled neurons on the ipsilateral side). On the ipsilateral side their is to observe an increasing of labelled cells in the rostro-caudal direction. Mainly in the caudo-dorsal region of the LC their are about 50% of the HRP-containing cells. The labelled cells belong to the medium-sized multipolar type, seldom they are fusiforme or ovoid. 2. In the NCS labelled cells are distributed throughout the whole nucleus but they are frequently to observe in the medium and caudal part near the midline. They belong to the multipolar and fusiforme cell type; labelled ovoid cells are rare. 3. The NRD contains labelled cells especially in the median and caudal part. The rostral part shows only a few HRP-containing cells. The labelled cells are mainly concentrated in the ventromedial NRD between the Fasciculi longitudinales mediales. The HRP-containing neurons are of multipolar, seldom of fusiforme or ovoid shape. In general, the number of labelled cells is higher in the LC than in the two raphe nuclei. After injection of HRP in the caudal part of the hippocampus or in the ventral hippocampus the number of labelled cells in the studied nuclei is lower than after injection into the rostral and medial part of the hippocampus. The results demonstrate that as well the noradrenergic as the serotonergic nuclei have an important connection with the hippocampus. The sources of this projection demonstrate a neurotopistic order. The functional importance of these connections is discussed.

The influence of orotic acid and sodium orotate on the differentiation of mitochondria of the neurons within explantate cultures of the hippocampus of 18 day old rat embryos was investigated by means of morphometrical and stereological methods. The ultrastructure of the mitochondria in controls and pharmacologically influenced cultures was analysed on electronmicrographs after 3, 13 and 20 days in vitro. According to their morphological appearance and state of preservation the mitochondria were classified in 3 types: juvenile mitochondria (stage I), active mitochondria (stage II) and destroyed mitochondria (stage III). After 3 days of cultivation the neurons in cultures treated with orotic acid and sodium orotate showed significantly higher rates in the numerical density of mitochondria (ratio of the number of all mitochondria per unit volume of cytoplasm) in comparison with controls. Moreover, sodium orotate treatment lead to a significant increase especially in the numerical density of the mitochondria of stage II (active mitochondria) on the third day in vitro. However, the mitochondrial volume per unit volume of cytoplasma of the neurons in drug influenced cultures is lower than in controls for each of these dates--3., 13., and 20. day in vitro. The volume of the mitochondria increased under treatment with sodium orotate from the 3. up to the 20. day of cultivation but never reached the values of the uninfluenced controls. The plastic changes of the mitochondria due to pharmacological treatment with RNA--precursors demonstrate the possibility of a stimulating effect of the drug on the differentiation of neuronal mitochondria in vitro.

Based on the findings of alterations in the chemoreactivity of neurons under conditioning and its possible significance for the formation of transient neuronal junctions, the authors investigated the influence of electrical stimulation of the Locus coeruleus upon the transmitter sensitivity of cortical and hippocampal neurons. The majority of neurons showed immediately after stimulation of the Locus coeruleus a different reaction to microiontophoretic application of acetylcholine, dopamine and glutamate than under normal conditions. Obviously there must have occurred qualitative changes because a superposition effect of the reactions following stimulation of the Locus coeruleus and application of the substances has rarely been observed. The release of noradrenaline provoked by the stimulation of the Locus coeruleus evidently leads to qualitative changes in the neurone texture, which may occur both inter- and intraneuronally.

In this paper qualitative investigations by means of light microscopy (Golgi impregnation) considering the ontogenesis of efferent neurons of the CA3, CA4a, CA4b regions in the rat hippocampus are presented following the already described development of CA1 pyramids (MINKWITZ and HOLZ 1975). After the migration of neuroblasts the types of projective neurons can be distinguished in the mentioned regions at the end of the prenatal period. During the first postnatal time particularly the basal dendritic tree develops expressing more distinctly the typical forms of pyramids (CA3, CA4a) and the of the multipolare neurons of CA4b. No distinct qualitative jumps at the transition of the regions in contrast to the CA3--CA1 boundary can be observed. Temporal retardations in the appearance of postsynaptic structures (microdendrites, excrescenses, spines) at pyramidal neurons from CA4 to CA3 and CA1 are regarded as signs of earlier development of the phylogenetic older pool of cells. During the early postnatal developmental period (until stage P10) proceedings of the growth are predominant. Later on, during the late postnatal period, the maturation of the neurons takes place with a considerable multiplication of spines distributed in similar ways as known from other pyramids. At stage P20 neurons of all hippocampal regions cannot be distinguished from adult ones without using morphometric procedures. The development of parts of the intrahippocampal network including mossy fibre system and SCHAFFER collaterals is discussed with regard to in this way appearing boundaries between fascia dentata and regio inferior CA4, CA3) and regio superior (CA1). These fibre systems establish by their stepwise construction one of the basic elements to hippocampal functioning.

In this paper pyramidal neurons of the CA 1-region of the hippocampus of the rat were investigated by means of GOLGI-methods. Animals were used in the stages of age E 20, P 5, P 10, P 15, P 20 added by 15 days old rats reared in darkness or under the influence of constant noise of 80 dB. In these cells the dendritic structure and the amount of visible spines were morphometrically estimated and evaluated by statistical procedures. The neurons of the normal ontogenetic range increase in dendritic length already from P 5 under particular emphasis on Stratum oriens and Stratum radiatum. The spines express a rapid development in the Strata oriens, radiatum, lacunosum beginning in Str. lacunosum at P 5 and in the others at P 10. This fact indicates certain systems of afferents to arrive at several times or to posess different importance to the layors. Because of the fact, that the Str. moleculare, which receives only extrahippocampal afferents, remains significantly behind the other layers regarding the development of spines, it is to be assumed, that the intrahippocampal afferents exhibit a higher importance to the development of CA 1-pyramids. The rearing in darkness causes a significantly decreased development of spines within all the layers, so that the amount of visible spines nearly remains at the values of P 10. Constant noise however causes only a significantly enforced development of spines in the Strata oriens, radiatum, molecular e.g. in layers receiving extrahippocampal afferents. An approach to stage P 20 takes place only at Str. moleculare possibly depending on its large extrahippocampal input. As an expression of specificity a pericaryon-dendrite-factor can be assumed responsible for the election of afferent fibres and the development of spines especially at the animals reared in darkness. The reactivity of the neurons is exhibited by the relative independence of the Strata lacunosum and oriens during the ontogenetic development and of the Stratum moleculare at rats reared under the influence of constant noise.

Following GOLGI impregnation pyramidal neurons of the rat hippocampus (CA1) from four stages of age (E21, P5, P10, P20) were investigated by morphometric methods with regard to their dendritic structure and amount and distribution of spines. The daesults show, that during the ontogenesis until reaching adult values the determined parameters are strongly following in their expression the sequence: degree of ramification, length of dendrites, amount of spines, and in the development of the certain dendritic parts of the pyramid the sequence: main apical dendrite with end ramifications, basal dendrites, apical lateral dendrites. Along the main apical dendrite the pattern of spine distribution develops proceeding from the form of a reverse nearly plain parabola (P5, P10) to the curve known from other adult pyramides (P20) with a spine poor initial segment, then the rapid increase in the number of spines (maximum of spine density in the middle of the whole distance), followed by a subsequent slow decrease in the number of spines. The apical end ramifications are mostly developed in the degree of ramification and length of dendrites during the prenatal period and show postnatal only significant changes in the spine density without reaching the values of the other dendritic fields. At basal dendrites no further increase in the number of dendrites can be found after P10. In dendritic length and spine density they are reaching adult level about the 20th postnatal day. In comparison with that, apical lateral dendrites show at P20 their definitive amount and a further increase of the spine density must be expected. The single dendritic fields of apical lateral dendrites are in length and total amount of spines sometimes smaller than those of basal dendrites, but more abundant. In both cases the spine densities increase until the stage P20 but they are not differing. Caused by the further development of apical lateral dendrites to adultness a predominance of the spine density of this part of the neuron is known. The total length of the dendrites of one pyramidal neuron (CA1) increases from 1000 mum (E21) to 5600 mum (P20), while the total amount of visible spines and the spine density increase from 20 to 3000 spines resp. from 0,02 to 0,55 spines/mum. It is important that the portion of the apical lateral dendrites grows from 2 to 40%, the portion of basal dendrites only from 20 to 33%, while the importance of the apical end ramifications decreases from 60 to 20%. In the presented paper the comparisons of developmental stages of the most pyramidalized hippocampal neurons (CA1) with adult pyramids of this region and with cortical pyramidal neurons of different phylogenetic ages are discussed.

The CA 1 pyramidal neurons of the dorsal and ventral hippocampus as also the neurons of the granular layer of the parietal cortex, which were investigated in comparison, show karyovolumetrically revealed significant variations of their activities during the postnatal development of the rat (between the 5th and 60th postnatal day). These variations represent the total expression of the gene activities related to the development and differentiation as also to the synchronously requested specific performances of the neurons for the organism. They are characterized by a more or less distinct regional specificity as also a sexual dimorphism--especially distinct during the first 3...4 weeks of postnatal development--beside several distinct principal coincidences. Characteristic are the peaks of activity at the 10th and 20th day of postnatal life and additionally at the 45th day in male animals. The activation of the hippocampal pyramidal neurons as also of the neurons in the granular layer of the parietal cortex from day 5 to day 10 of the postnatal development, which comes to appearance as an increase of cell nuclear volume between 35,2 and 142,5 per cent, coincides with the growing period of the central nervous system, which lasts in the rat till the 10th postnatal day, and with the postnatal initial phase of several extracerebral developmental, differentiation and maturation processes. The variations of neuronal activities occurring after the 10th day and ceasing in the 4th week of the postnatal development coincide with the maturation period of the central nervous system and of the extracerebral systems. The peak of activity appearing around the 45th day of life in male rats seems to be related to the puberty. The sexual dimorphism of the hippocampal pyramidal neurons as also of the neurons in the granular layer of the parietal cortex, realized in the cell nuclear volume resp. neuronal activity, is apparently the reflection of the sexual specific differentiation of the central nervous system and is a partial phenomenon of the complex dimorphism of the both sexes. It shows also a regional specific character.

In the vicinity of the collateral sulcus the cellular islands of the entorhinal region (lamina alpha of the outer principal layer = Pre-alpha) fuse, forming a cellular plate which runs obliquely through the outer laminae. Finally, the cellular elements of Pre-alpha lie in between the third and the fourth layer of the isocortex. The islands are mainly composed of star-shaped nerve cells with thorny dendrites and an axon extending into the white matter. Within the reaches of the oblique plate the shape of these cellular elements underlies an alteration. Apical and basal dendrites become more and more recognizable, the cell body gains the shape of a pyramid. For this reason, we consider the star-shaped neurons of the islands to be modified pyramidal cells. They are compared with the genuine star cells (Golgi-II-cells) of the layer. Distinguishing characteristics not only of the Golgi- but also of the pigment-picture allow the unequivocal distinction between the modified pyramids and the Golgi-II-cells.

1. The development of pyramidal neurons of the hippocampus (CA1) of the albino rat was investigated by means of light-microscopy with GOLGI impregnation methods. The stages of age of the animals covered the period from the 15th prenatal to the 30th prematal day. Three and six months old rats were used for comparison. In addition some histological series were made by staining with hemalaun-eosine. 2. The formation of the apical, basal and side dendrites, the existence of common signs of growth (swellings, growth cones, sprouts) and the amount of spines were used for the estimation of neuronal changes. The development of the typical structure of the hippocampal region and of the pyramidal layer in the CA1-field was described. 3. The migration of neuronal elements to the pyramidal layer is finished around the time of birth, but lasts in the area dentata at least to the 8th postnatal day. The hippocampus shows a nearly mature figure in its hitology 15 days after birth. 4. Until the 19th embryonal day bipolare spongioblasts can be seen, but already one day before postmigratory neuroblasts are existent. During this period the division of matrix cells occurs and the migration of the neuroblasts is beginning. (early prenatal developmental period) 5. Since the 19th prenatal day the neuroblasts show long apical dendrites, growing axons with first sprouts of collaterals, but also sprouts of basal and side dendrites. Until the time of birth a further formation of dendrites and a narrowing of the perikarya in the pyramidal layer follow. (late prenatal developmental period) 6. The intermediate neuroblasts grow continuously in the first postnatal time and let perceive an increase in the number of dendritic stems and branches. Postsynaptic structures (spines) appear in a little increasing number. The initial segment of the apical dendrite remains free of spines. (early postnatal developmental period).

The regional distribution and cellular localization of aspartate aminotransferase (AspAT) as a glutamate and aspartate metabolizing enzyme was studied in the hippocampal formation, in dorsal root ganglia, and in superior cervical ganglia during the postnatal development of the rat. At birth, in all neuronal tissues studied the enzyme activity was rather low. Whereas in the dendritic layers of the hippocampal formation enzyme levels rose strikingly during the first weeks of postnatal life, those of peripheral ganglia remained remarkably stable. The high affinity uptake of [3H]labelled L-glutamate and D-aspartate were studied in surface autoradiograms of incubated slices of hippocampal formation during postnatal development of the rat. Our data indicate that the postnatal development of high affinity uptake capacity is parallel to the increase of histochemically demonstrable AspAT activity in the hippocampal formation suggesting that the enzyme may be involved in glutamate/aspartate neurotransmitter metabolism.

Using the E-PTA procedure of Aghajanian and Bloom we determined the numerical density of synapses in the stratum lacunosun-modeculare of the CA1-region in the hippocampus of male albino rats. On this occasion the sources of error both the natural and the methodical were investigated. As natural causes of remarkable scattering and failing correspondence in the findings the individual variability of the synaptic density and the inhomogeneous distribution of synapses must be taken into consideration. Concerning the methodical sources of error the differences in tissue shrinkage become apparent. But faults resulting from applying morphometric methods are the most serious. The counting of synaptic profiles is based on defined morphological criteria. The portion of the synaptic profiles recorded in this way in relation to all those present in the section depends beside other things on the section thickness. Accordingly the correction factors are in correlation with section thickness. The application of the Floderus correction presupposes the determination of the minimum length of the counted profiles. As a result of these investigated the possibilities for reducing the error have been demonstrated.

1. The influence of the cyclic N6-monobutyryladenosine-3':5'-monophosphate (mb-cAMP) on the neurogenesis was investigated by means of electron microscopic and morphometric methods at the explantate cultures of the fetal hippocampus of the rats in vitro. 2. After three days of the in vitro cultivation the neuroblasts of the treated cultures show not significantly changes of the total number of ribosomes in comparison to the controls. 3. After 7 and 14 days of the in vitro cultivation the neurogenesis is significantly stimulated under the influence of the mb-cAMP. The drug produces a significant increase of the free ribosomes and polysomes in company with a higher degree of differentiation of the granular endoplasmatic reticulum. Under the influence of the mb-cAMP the total number of ribosomes significantly increases by 32% (after 7 days) respectively 20% (after 14 days). 4. The results might suggest, that mb-cAMP might stimulate the protein synthesis in connection with an increased nerve fibre growth in vitro.

150 hippocampal explants from fetal and neonatal rats were cultivated with semisynthetic media in maximow chambers without movement and in roller tubes by rotation in the virutor up to 3 days. The cultures were fixed in Bouin, cuted 15 microns and coloured with the method of Klüver-Barrera or haemalaun. Histological examination of the explants revealed that hippocampi cultured in maximow chambers without movement possess an organisation similar to that observed in situ. Experiments with rotated explants of hippocampus indicate that the structure of hippocampus in vitro is changed. Especially neurons are degenerated and the migration of cells is altered. Finally the value of these two methodological cultivations for neurobiology is discussed.

Explants of hippocampus from fetal rats were cultivated in Maximow chambers in semisynthetic medium up to 12 days in vitro. The cultures were fixed Bouin, slided 15 micron, coloured with Klüver-Barrera and some morphological parameters were tested. 1. The nerve fiber index increased by influence of 1% Solcoseryl in relation to control cultures, which growed in minimal medium. An essential stimulation was observed by application of placentar serum and embryonal extract into the culture medium. 2. Die decrease of the number of neurons and glial cells per unit of area and a small decrease of the area of neuron nuclei was discussed in relation to the effect of the pharmacon Solcoseryl on O2- consumption. 3. Solcoseryl (firm Solco AG, Base) is an extract of calf blood. It can not substitute other tissue extracts.

A total of 60 newborn Wistar-rats was divided into three groups, each of which was exposed to a different environmental situation. The first group was reared in complete darkness while the second one was subjected to permanent noise. The third group (controls) was reared under normal laboratory conditions. The animals of all three groups were sacrificed on postnatal day 15 and prepared according to the Golgi-Kopsch technique. Apical oblique dendrites and basal dendrites of hippocampal pyramidal cells (CA 1 region) were ranked in a centrifugal ordering system for measurements of dendritic length and for spine counts. In the successive dendritic orders as well as in the dendritic field (all branches of a dendrite arising from the cell body or main apical shaft) no statistically significant differences between the groups were found with regard to dendritic length. The spine counts, however, revealed significant changes. In the dark reared group, the number of spines on oblique dendrites as well as on basal dendrites is significantly decreased in comparison with the controls. On the contrary, animals subjected to permanent noise exhibit significantly more spines on oblique and basal dendrites when compared with the controls (mean number of spines in the apical dendritic field: controls 30.1 +/- 26.9, dark reared animals 17l1 +/- 15.4 (-43.2%), animals subjected to noise 36.4 +/- 31.4 (+ 20.9%); basal dendritic field: controls 82.2 +/- 70.1, dark reared animals 47.3 +/- 35.1 (-42.5%), animals exposed to noise 114,1 +/- 94.6 (+ 38.8%)). These results confirm previous studies on the number of spines along the main apical shaft of CA 1 pyramidal cells (Frotscher et al. 1975) and on the number of synapses in the stratum radiatum of CA 1 (Frotscher et al. 1977) in rats reared under identical experimental conditions. On the basis of these results it seems to be obvious that synaptogenesis in the hippocampus is influenced by the environmental conditions used though this region is not directly involved in the optic or in the acoustic pathway.

In this paper qualitative results are presented from GOLGI preparations by light microscopic evaluation about the appearance and the ontogenetic changes on GOLGI-II-type neurons of the rat hippocampus from 15 stages of age between the 20th embryonic and the 20th postnatal day. Interneurons are already detectable in all the hippocampal regions in the prenatal period. Their postnatal growth expresses above all in lengthening rather than in multiplication of the amount of dendrites. Cells predetermined for developing spines show already early in the postnatal period a rather large amount of spines. The further multiplication of the spine density remains minimal that never values of adult pyramids will be reached. The form of the dendritic and axonic fields of neurons with short axons is very variable. Nevertheless three main categories of GOLGI-II-type neurons can be distinguished: 1. neurons, the axonic and dendritic ramifications of which strongly remain within the layer (do not exist within the stratum pyramidale), 2. neurons, which are able to contact afferent fibres in other layers by means of ascending and/or descending dendrites, 3. basket cells, the axons of which have direct influence on the perikarya of pyramidal neurons. Within the region CA4b interneurons of the 2nd group could be found, dendrites of which penetrate into layers of the fascia dentata. Such a fact does not exist at the multipolare, efferent neurons of the same region and could never be seen at any axonic ramification. In the presented paper aspects of influencing the maturation of pyramides by interneurons are discussed. Also has been tried to summarize the results about the neuronal structure of the rat hippocampus by schemes of frontal sections from four selected stages of age.

Male albino rats aged 13 weeks experienced a single training for bright -- dark discrimination in a Y-chamber. The numerical density of synapses determined in the stratum lacunosum-moleculare of the hippocampal area CA1 24 hours later turned out to be lower by 3.4 per cent with trained animals compared to passive controls. This difference failed, however, to prove significant.

In the brains of male white rats some structural parameters of granule neurons in the hippocampal dentate gyrus such as cytoarchitectonical arrangement of these cells, structure of dendrites and dendritic spines were analyzed during ontogenetic development as well as changes in these parameters due to modified external conditions (acoustic stimulation, dark rearing, cerebrolysine administration). Transmission electron microscopy as well as cytologic and morphometric methods were employed. Under normal conditions the adult cytoarchitectonical pattern of the dentate gyrus was reached at postnatal day 20 (P 20). At the ultrastructural level the differentiation of the neuronal perikarya during ontogenesis was determined by the size of the nucleus, occurrence of rough endoplasmic reticulum, ribosoms and polysoms, Golgi apparatus and mitochondria between birth and postnatal day 90 (P 0--P 90). Synaptogenesis starts in the perinatal period in form of immature axo-somatic and axo-dendritic contacts. In adults a pronounced structural variability of synapses and the presence of complex spines was found. The average density of spines increased considerably with age in isolated dendritic segments as well as over the whole length of a dendrite. The density was at P--P 2: 0,03; P 5: 0,04; P 8: 0,09; P 10: 0,1; P 15: 0,17; P 20: 0,23; P 30: 0,48; P 180: 0,62 spines per micrometer dendrite. The postnatal development of the neurons under experimental conditions was characterized by significant augmentations of the spine numbers mainly in the early postnatal phase: Controls at P 15 with 0,17, after dark rearing 0,30, after Cerebrolysine 0,32, after acoustic stimulation 0,33 spines per micrometer dendrite. At P 30 only dark rearing resulted in significant augmentation of the spine number (0,63 spines per micrometer dendrite against 0,48 in controls). The data obtained after acoustic stimulation and dark rearing point to a widespread influence of external stimuli on neuronal development not only confined to the corresponding visual or acoustic pathway. The Cerebrolysine results during early postnatal development can be explained by a stimulation of protein synthesis and, hence, of the neurogenesis.

The dendritic spines of the apical and basal dendrites of pyramidal neurons in CA1 and CA3 and of granule cells in Fascia Dentata were investigated in 19 stages of age, from the 10th to the 190th postnatal day. The results were statistically evaluated. In CA1 and CA3 a first increase in the number of spines (with a maximum at 35 days) is followed by a subsequent decrease up to the 65th day and a further increase up to the 190th day. This developmental pattern is found in the apical dendrites of CA1 and CA3 in all its investigated segments. On the contrary the basal dendrites differ from this pattern. The spines in the Fascia Dentata show a rapid increase up to the 15th postnatal day followed by a slow but constant increase up the end of our study.

Organ cultures of hippocampus from fetal rats were cultivated in Maximow chambers with semisynthetic media up to 12 days in vitro and fixed after 1, 5 and 12 days in culture, cuted 15 micrometer and coloured with the method of Klüver-Barrera. The effect of 5 x 10(-6) M sodiumorotate and 6 x 10(-6) M orotic acid was tested. 1. The density of cells is related to the zone of cells, the drug and time of incubation. 2. The density of cells increased in the zone of matrix after application of sodiumorotate and orotic acid. The density of cells significantly decreased in the migration zone. The density of neurons significantly increased in the region CA 3. 3. Orotic acid and sodiumorotate influenced the migration of cells in vitro. Sodiumorotate stimulated migration and differentiation. Longtime cultured hippocampi degenerated in vitro after influence of orotic acid. 4. The area of nuclei from neurons increased up to 12 days in vitro, when the cultures were tested with sodiumorotate. Only within the first 24 hours orotic acid effected an increase of the area of nuclei from matrix cells. It is discussed, sodiumorotate influenced in vitro metabolic processes in suitable systems.

Caryovolumetric investigation of hippocampal neurons performed in intact female rats at 21, 26, 32 and 36 days of age, and on the day of the first vaginal estrus revealed distinct prepubertal changes of nuclear volumes of pyramidal neurons located in the ventral hippocampus close by the subiculum. The caryovolumetrically demonstrable activity of these neurons increased significantly between days 21 and 26 and showed a subsequent decrease up to day 36. It was again significantly enhanced at the time of the first vaginal estrus. In contrast to the ventral hippocampus, neuronal activity of the dorsal hippocampus was at a constant level throughout the prepubertal development. Neonatal ovariectomy reduced significantly the nuclear volumes in both hippocampal regions and, simultaneously, prevented completely the prepubertal fluctuation of neuronal activity recorded in the ventral hippocampus of intact rats. The results suggest that the prepubertal activity of hippocampal neurons depends on the integrity of the cerebro-hypophyseo-gonadal axis. They furthermore support the conclusion derived from experimental studies that the hippocampus may be involved in the neurohormonal control of female puberty.

The neuronal structure and synaptology of the hippocampus of the rat are studied using neurohistological, electron microscopical and morphometrical methods. After Golgi-impregnations the neurons of the hippocampus were classified by reason of special morphological features (pericaryon, dendritic, ramification, course and termination of the axons). The neurons of the hippocampus show from the region CA4 towards the region CA1 an increasing "pyramidalization" which is explained respecting the neuronal structure and synaptic architectonics as the result of a specific presynaptic representation of afferents at the neurons. The high degree of differentiation of the CA1-pyramids speaks for the progressive developmental tendency in the phylogenesis of these pyramids compared to the neocortical neurons. The ultrastructural investigations of the hippocampus show that each individual layer of the regions CA1, CA3 and CA4 has a characteristic synaptic architectonics. In all hippocampal layers the axo-spino-dendritic contacts are the most frequent type of synapses with the structural characteristics of the excitatory synapses; the synaptic contact appears as "Bouton en passage" respectively as "Bouton terminal". The axosomatic synapses of the stratum pyramidale (CA1 to CA3) predominantly are developed as symmetrical contacts with inhibitory function. As the terminals of the basket cells they represent the morphological correlate for the recurrent inhibitation of the pyramids by the inhibitory effecting basket cells. In the supra- and infrapyramidal layers of the regions CA3 and CA4 special contacts are developed for the connection withe the mossy fiber terminals--at the pericarya and the main dendrites as so-called somatic and dendritic protrusions and as micro-dendrites. As has been established for the neocortex, the dendritic spines are the decisive receptive postsynaptic structural characteristic of the hippocampus as well. Pre- and postsynaptic elements show a broad scope of variation in forming their structures. This also applies to the intrahippocampal systems with their highly complex structure which is the morphological expression of an extremely high degree of differentiation. The morphometrical investigations of the ultrastructure of synapses enclose the calculation of the number of synapses per mm3 of the volume of layers, the average individual volume of synapses, the presynaptic portion of volume of the relative volume of cortex of the hippocampal layers as well as the average number and size of the synaptic vesicles. The average number of synapses per mm3 brain tissue runs in CA1 to 5,9 X 10(9), in CA3 to 7,9 X 10(9) andin CA4 to 8,7 X 10(9) synapses per mm3. The average individual volume of synapses amounts to 0,2-0,3 mm3 whereas the average portion of volume of the presynaptic structures per relative cortical volume comes to 14% in CA1, 16-19% in CA3 and 15% in CA4.

Explants and cell cultures of embryonic chick ganglia trigeminalia, telencephalon and retina or hippocampus from fetal rats were incubated in maximow chambers in the presence of cyclic AMP and the dipeptide cyclo(Lys-Pro).HCL under various conditions. Maintenance of nerve cells and growth of nerve fibers were observed by morphometrical methods. 1. Cyclo(Lys-Pro).HCL promoted the maintenance of neuroblasts and the growth of nerve fibres in explants of the ganglion trigeminal and retinal cell cultures. The effect depended on the presence of serum in the medium by use of poly-I-lysine substrate. 2. Extern applicated cyclic AMP and the dipeptide SP3-4 = cyclo(Lys-Pro).HCL facilitated neurite growth in PNS cultures. In the presence of the drugs the length of nerve fibers increased for a short term. On CNS explants substance P (SP1-11) and SP3-4 were without effect. Cyclic AMP stimulated the growth of nerve fibers in CNS explants and cell cultures in number and length. 3. Discussed is the effect of SP1-11 and cyclo(Lys-Pro).HCL for competence of nerve fibre regeneration in vitro in relation to increasing cAMP levels, which may then act as an initial second messenger. It is suggested that explants and cell cultures of nervous tissues will be useful as a tool for the further characterisation of factors with neuronotrophic activities.

Explants of nervous tissue (CNS: hippocampus from fetal rats, telencephalon and PNS: ganglion trigeminale from chick embryos) were cultivated in maximow chambers. Biological extracts (brain and embryo extract) or chemical agents (aminoacid mixtures and peptides with known sequence of amino acids) were tested. Biological extracts and aminoacid mixtures stimulated the differentiation in PNS and CNS cultures. A stimulating effect of peptides seems to exist only on PNS explants. The fundamental importance of suitable reference systems, parameters, optimal concentrations and periods of application of effective substances for the evaluation of the results is discussed.

Explants from chick embryo PNS (ganglion trigeminale) and from CNS of embryonal rats (hippocampus) and dissociated cells from chick embryo cerebral hemispheres were cultivated in maximow chambers in the presence of various concentrations of placental serum and of a chemically synthesized tripeptide Gly-His-Lys. 1. The presence of tripeptide in the nutrient medium with a low concentration of serum did not compensate the outgrowth of nerve fibers, that take place in the growth medium. 2. In the presence of tripeptide in the nutrient medium with low concentration of serum the index of growth area increased significantly. 3. Within the first days in cell cultures 0,01 microgram tripeptide pro ml medium stimulated the outgrowth of neuronal processes. 4. The experiments indicated, that the tripeptide did not replace the serum. The possible role of tripeptide as a system in controlling neuron-glial ratio in vitro is discussed.

Electron microscopical results on "special structural relations" between neighbouring neuronal elements (pericarya, dendrites) are demonstrated in different regions of the rat limbic system: field CA3 and field CA1 of the hippocamp, dentate gyrus, area entorhinalis (L V) and area retrosplenialis granularis (L III). The "special structural relations" are to be characterized as follows: direct apposition of membranes of the neighbouring neurons including an extracellular space of relatively constant width over a larger distance without neuroglia intervening this space. Areas of membrane appositions have been observed between neuronal somata ("soma-somatic appositions"), between dendrites ("dendro-dendritic appositions") and between both, somata and dendrites ("dendro-somatic appositions"). In the elmigraphs their length varied between 1 and 6 microns. In the entorhinal cortex and in the retrosplenial cortex soma-somatic appositions were in the majority, whereas in the hippocampal regions dendrodendritic appositions, in the dentate gyrus all types seem to be predominant. In the literature such direct appositions of neuronal membrane often have been described in the CNS of different species. They are considered as a possible morphological correlate of the ephaptic interaction, which is caused by field effects between neighbouring neurons. Following the literature the results of the ephaptic interactions vary between a weak facilitation and synchronous discharge of an unstimulated neuron by a stimulated one. Although such direct appositions are present in different groups of neurons in the limbic system, especially in the hippocamp, up to now their functional meaning is unclear. No correlation seem to exist between neuronal membrane appositions/ephaptic interactions and such a phenomenon like the hippocampal longterm potentiation; mechanisms of cooperativity included in the LTP like coactivation effects of pre- or postsynaptic elements still remain to be clarified.

Cell cultures from hippocampus of 16 and 17 days old embryonal rats were cultivated up to 4 weeks. After 24 hours in vitro on 18.4 percent of cells and after 5 days in vitro on 70 percent of cells processes could be recognized. These are neuroblasts. The cells reaggregated. Nerve fibers after 4 weeks in vitro are 200 to 300 mum long. Small and big neurons with 12 mum to 26 mum diameters of perikarya, bi- and multipolar neurons after 4 weeks in vitro were observed. In cultures and meningothel-monolayer developed. Maintenance and differentiation of cultures are possible only by sowing in at least 60,000 cells/ml medium. The advantage of cell culture opposite to organ culture exists in experiments with immediate selective influence.

Explants of the hippocampus (rat) were cultivated either in maximow chambers without movement (static) or in movement (dynamic) by rotation in roller tubes. The growth of nerve fibres, the density of cells (matrix, neurons) and the area of cell nuclei was observed. The determination of the index of nerve fibre growth, the procedure for counting cells in a tissue with different densities and the value of the two methodological approaches of cultivation is discussed. Finally it is stated that best results in morphology of explants can only be obtained by cultivation without movement.

In the way of quantitative investigations of nerve cells and neuroglia in g. parahippocampalis and following analysis of measuring dates by multivariat variance- and discriminance-analysis was demonstrated that aging process in ammonite formation can be described on the base of cells only in sector subiculum.

The entorhinal afferences to Fascia dentata and Stratum lacunoso-moleculare of CA1 could be visualised by means of anterograde axoplasmic transport 5 days after L-[5-3H]-proline injection into the entorhinal cortex. Furthermore, the macromolecule-bound tracer was found in the perikarya of granule cells and mossy fibres as well as in the cell bodies of the CA1 pyramids. These findings indicate that labelled material, released from the axonal terminals of the perforant pathway, is taken up by these neurons.