Surface structure of nerve tissue transplants growing in the anterior chamber of the rat eye (electron-microscopic study)

Two different types of the glial envelope are demonstrated around the septal cerebral area grafts (SCAG) and the hippocampal grafts (HG) developing in the anterior eye chamber of the rat during 3-4 months. In the SCAG it is multilayered, consists mainly of modified oligodendrocytes. Processes of the nervous cells perforate the surface of the envelope. In the HG the unilayered glial envelope consists of bodies and processes of the fibrous astrocytes; it is impenetrable for the nervous processes. Since both types of the cellular elements making these envelopes have been described in the norm, it is possible that the differences in the surfaces described are specific for these structures.     

Transplantation immunology of the anterior chamber of the eye. II. Immune response to allogeneic cells.

The mechanism by which the anterior chamber of the eye extends immunologic privilege to allogeneic donor tissues has been studied in inbred rats. Inoculation of allogeneic lymphoid cells into the anterior chamber demonstrated that although the site lacks a lymphatic drainage, the afferent limb of the immunologic reflex arc is intact because the recipient can recognize and mount a specific immune response. In addition, host immunity was able to express itself within the anterior chamber when induced systemically, indicating that the efferent limb of the reflex arc is also intact. Therefore, it is suggested that the unique immunologic features of the anterior chamber may result from the obligate intravenous presentation of graft antigen to the host's systemic immunologic apparatus, a route that prejudices the host's response in the direction of tolerance and/or enhancement rather than cell-mediated, tissue destructive immunity.     

Embryologic development of rat adrenal medulla in transplants to the anterior chamber of the eye

The morphological development and plasticity of embryonic and postnatal rat adrenal medullary cells were studied in homologous adrenal grafts to the anterior chamber of the eye. The eyes of recipient rats were adrenergically denervated 10 days prior to grafting by extirpation of the superior cervical ganglion in order to increase levels of NGF and NGF-like activities in the iris. Grafts taken at the 15th day of embryonic development (E15), i.e., at the beginning of immigration of medullary progenitor cells into the adrenal cortical anlagen, contained no cortical or mature medullary cells after 2 weeks in oculo. Numerous sympathoblastic cells, however, were located at the anterior surface of the iris. E 16 and E 17 transplants showed abundant mature cortical tissue after 2 weeks. Small groups of medullary cells with the ultrastructural characteristics of mature pheochromoblasts or young chromaffin cells were interspersed among cortical cells without forming a discrete medulla. Neuronal cells were exclusively found outside the cortical cell mass. Sympathoblasts grew at the surface of the iris, while young sympathetic nerve cells, which were invested by Schwann cells and received synaptic axon terminals, were embedded into the stroma of the iris. Grafting of E 21 adrenals yielded very similar results except that, in a few instances, young chromaffin cells were located outside the cortex and sympathetic nerve cells were seen to be in close contact with cortical cells. In transplants of adult medullary cells typical mature adrenaline and noradrenaline cells were clearly distinguishable after 8 weeks even in the absence of cortical cells. The only indication of phenotypical changes in these cells was the formation by some of them, of neuritic processes which could be visualized in glyoxylic acid-treated whole mounts of irises. These results are compatible with the idea that embryonic adrenal medullary cells have the environmentally controlled potential to develop along the neuronal or endocrine line, but could also be interpreted in terms of a selection of a specific subpopulation with predetermined potentialities by a specific microenvironment. Moreover, these results suggest that increasing differentiation of medullary cells is accompanied by progressive restrictions in their genetic program, which eventually prevent full transdifferentiation of mature chromaffin into neuronal cells.     

Insulin-like growth factors decrease catecholamine content in PC12 rat pheochromocytoma cells.

Insulin-like growth factors (IGFs) stimulate proliferation and differentiation of PC12 rat pheochromocytoma cells and modulate catecholamine release in bovine adrenal medullary cells. Dexamethasone increases catecholamine synthesis in PC12 cells. We therefore studied the effects of IGFs and dexamethasone on catecholamine content in PC12 cells. Dopamine (DA) and norepinephrine (NE) content of PC12 cells were measured after incubation for 72 h with IGFs (100 ng/ml) and/or dexamethasone (500 nM). IGF-I (100 ng/ml) and IGF-II (100 ng/ml) decreased DA and NE content to approximately 35% and approximately 25% of control, respectively. [Leu27]IGF-II, which binds to the IGF-I receptor with markedly decreased affinity, did not reduce catecholamine levels, indicating that the effect is likely to be mediated by the IGF-I receptor. Dexamethasone (500 nM) increased levels of DA and NE to 173 +/- 20% and 331 +/- 48% of controls, respectively. Coincubation with IGFs did not significantly affect the stimulation of DA by dexamethasone, but abolished the rise in NE. Levels of tyrosine hydroxylase mRNA, protein and activity were increased following incubation with dexamethasone, but were unchanged by IGFs. These results indicate that IGFs decrease catecholamine content in PC12 cells via the IGF-I receptor. Complex regulation involving multiple synthetic and/or degradative steps is implicated in this process.     

Post-translational processing and secretory pathway of human atriopeptin in rat pheochromocytoma cells.

Atriopeptin (AP) is expressed in several tissues with each tissue capable of specific differences in processing of the prohormone (pro-AP) to mature low molecular forms of the peptide. Since pro-AP has low biological activity, processing into mature AP is a critical activation event. This observation prompted us to study whether granule storage or regulated secretion of AP is essential for cleavage of mature peptide. We examined the processing of AP in adrenal medulla derived cells, using the rat pheochromocytoma cell line (PC12 cell) stably transfected with a genomic human AP DNA in the presence and absence of nerve growth factor (NGF), and also examined the mechanism of AP secretion and compared the results with those obtained using transfected chinese hamster ovary cells (CHO cells). The amount of prohormone was 5-10 fold higher than that of low molecular form of AP in the transfected PC12 cells. This ratio was essentially unchanged in differentiated PC12 cells after NGF treatment of the cells. Potassium depolarization of the transfected PC12 cells caused a 5-fold increase in AP release into the medium primarily as the intact prohormone. On the other hand, transfected CHO cells only exhibited constitutive AP release which is non-response to depolarization. These results suggest that the AP prohormone is sorted into secretory granules as the prohormone in PC12 cells and undergoes regulated release in response to depolarization indicating granule storage or release is not the critical determinant of AP prohormone cleavage.     

Biogenic amine metabolites in human lung tumor cells: histochemical and mass-spectrographic demonstration

Three continuous cell lines isolated from small cell carcinoma of the lung (SCCL) have been examined for their ability to take up and metabolize the biogenic amine precursors, L-DOPA and 5-hydroxytryptophane (5-HTP). In two of the three lines formaldehyde-induced fluorescence (FIF) and mass-spectrographic analysis indicated specific uptake and metabolism of L-DOPA and 5-HTP.     

Electron microscopical study of synaptic glomeruli in cerebellum transplanted to the anterior eye chamber

Fetal cerebellar anlage from rat fetuses of 15-16 operational days were grafted into the anterior chamber of the eye of adult female albino rat recipients. Survival time of the transplants--containing both cerebellar cortex and cerebellar nuclei--was 2 to 2 1/2 months. Electron microscopical (EM) studies of the thin, under-developed granular layer of the laminated cerebellar cortex revealed the presence of well differentiated cerebellar glomeruli, surrounded by granule cell perikarya. As in the normal cerebellar cortex, the central profile of the glomerular complex was the large mossy terminal, containing spheroid synaptic vesicles, and forming synaptic contacts with dendrites and dendritic digits of the granule cells. Golgi cell axonal varicosities, containing ovoid or pleomorphic synaptic vesicles were found also on the periphery of the glomeruli. In addition, in several synaptic glomeruli, a third neuronal element was also observed, containing flat, discoidal vesicles and receiving synaptic contacts from mossy and Golgi axons, but being also presynaptic to granule cell dendrites. It is suggested that all mossy terminals in the cerebellar transplant originate from the cerebellar nucleus. Morphological evidence is also provided that the presynaptic dendrite-like processes--never found in normal cerebellar cortex--are also processes of nuclear neurons.     

Anterior chamber of the eye. A genetic and anthropological study in Greenland Eskimos.

During a population survey in Eskimos family resemblance with respect to anterior chamber depth (ACD) was studied. Heritability estimates of about 0.7 against a marital correlation of zero were obtained. No correlation between ACD and parity, but a significant correlation with stature was found which possibly accounts for the sex difference in ACD. Significant associations between ACD, iris colour, facial appearance, and corneal diameter were observed. West Greenlanders with a predominantly Eskimoan physiognomy showed smaller anterior chambers than unmixed East Greenland Eskimos and Eskimo-Caucasian hybrids.     

Change in the karyotypic structure of mouse and rat rhabdomyosarcomas on their transplantation into the anterior chamber of the eye

A study has been made of 7 transplatable lines of mice rhabdomyosarcomas and one line of rat rhabdomyosarcoma during their transplantation into the eye anterior chamber subcutaneous tissue. In all, 10 subcutaneous transplants and 15 transplants into the eye anterior chamber (EAC) were examined. Etanol fixed print smears were subjected to the Feulgen reaction to measure the DNA content using a cytophotometer MCPhU-1; 100 cells being measured in each transplant. In the majority of the EAC transplants, a statistically significant decrease of the karyotypic variability was found in additionto the augmentation to the diploid cell ratio as compared to subcutaneously proliferating populations of the same tumour lines. In some cases EAC transplants displayed exclusively diploid (periploid) populations of tumour myoblasts. Shifts in the karyotypic structure of populations towards diploidy, revealed during the cultivation of transplantable rhabdomyosarcomas, may be regarded as a phenomenon of the "karyotypical normalization" of tumour cells. The disappearance or sharp decrease of tetraploid or hypertetraploid classes of cells in EAC transplants may be due to the increase of their selective value in condition of immunological privilege of diploid, karyotypically normal cells, and of reduction of the genome mutation frequency in a diploid fraction of tumor myoblast populations.     

Growth of a graft from the septal region of the brain in the anterior chamber of the eye

The neural tissue (the hippocamp and septal cerebral area) obtained from rat embryos is transplanted into the anterior eye chamber of mature animals. The developing grafts are sized and measured and light optic microscopy is performed 3-4 months after transplantation. Essential differences in development and organization of the septal cerebral area (SCAG) and the hippocampal (HG) grafts are revealed. The SCAG have a spherical form, a limited contact with the host iris and are vascularized by one--two ingrowing and branching vessels. The HG are elongated in their form, have contacts with the host iris by the wide plane parallel to their long axis and a great number of blood vessels growing into them. The size of the HG is much larger than that of the SCAG. The latter contain diffusely scattered multipolar neurons, while the HG neurons are organized as a layer with their apical dendrites oriented towards the iris.     

Nature of macrophages in rat brain. A histochemical study

The nature of phagocytes appearing in lesions of the central nervous system is strongly debated with a tendency to assess an exclusively hematogenous origin. We studied the origin of phagocytes appearing in a stab wound in the rat brain. Histochemical stains for acid phosphatase and peroxidase, and silver impregnation techniques were used for our study. The results obtained showed the existence of two macrophage types: endogenous microgliocytes and exogenous monocytes.     

Immunoactive atrial natriuretic peptide in the rat eye: molecular forms in anterior uvea and retina

Acid extracts of rat anterior uvea and retina contain immunoactive atrial natriuretic peptide. Gel filtration chromatography demonstrates that the major forms of the uveal and retinal material have approximate molecular weights of 2400 and 1750 daltons respectively, similar in size to the hypothalamic form of the peptide but clearly distinguishable from the larger, cardiac form. When further analyzed by RP-HPLC, the immunoactive material from both ocular tissues elutes at a position similar to atrial natriuretic peptide-28, again distinguishing the ocular and cardiac forms of immunoactive material. These results suggest potential roles for atrial natriuretic peptide both in ocular vegetative physiology and in retinal function.     

Ultrastructure of nervous tissue developing in the anterior chamber of the eye. 1. Perikaryons and dendrites

Embryonic tissues of septum and hippocampus were transplanted into the anterior eye chamber (AEC) of adult rats. The morphology of initial embryonic tissues and of transplants within 3 to 4 months of cultivation in AEC was studied. The transplanted tissue consists of neuroblasts and immature neurones: no synaptic contacts are observed. Within 3 to 4 months, highly differentiated neurones establishing synaptic contacts can be seen in the transplants. At the same time the fine structure of perikaryons and dendrites undergoes some changes: increased vacuolization, transformation of ergastoplasm into lamellar bodies. These can be due to an elevated functional activity of some neurones. Another group of morphological abnormalities (increased number of dendrite processes and microphyllopodia, somatic spines, dendrite cones of growth, tight junctions between perikaryons) suggests incomplete tissue maturation. These might be due to the absence of normal afferent and trophic influences in AEC.