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Survival of embryonic dopamine (DA) neurons is extremely low (5-20%) following transplantation. Strategies to increase this survival are critical to the future of transplantation for Parkinson's disease. We demonstrate here that a factor(s) released from striatal oligodendrocyte-type 2 astrocytes (SO2A) greatly improves the survival and phenotype expression of mesencephalic DA neurons in culture while simultaneously decreasing the presence of apoptotic nuclear profiles, as detected by the TUNEL method and bisbenzamide/tyrosine hydroxylase double labeling. This SO2A-derived trophic factor(s) has minimal effects on glia and no effect on nondopaminergic mesencephalic neurons. The developmental period during which this SO2A trophic effect occurs (E14-18) coincides with the period when mesencephalic grafts are undergoing the highest rates of apoptosis, i.e., immediately following implantation. Therefore, SO2A-derived trophic factor(s) offers great potential for the augmentation of grafted DA neuron survival.

To enhance the current therapeutic benefit of dopamine (DA) neuron grafts in Parkinson's disease, strategies must be developed that increase both DA neuron survival and fiber outgrowth into the denervated striatum. Previous work in our laboratory has demonstrated that dopaminergic neurons grow to greater size when co-grafted with striatal cell suspensions and display extensive tyrosine hydroxylase-positive (TH+) projections, but no conclusion could be reached concerning enhancement of survival of grafted DA neurons. The aim of the present study was to characterize further the potential trophic effects of striatal co-grafts on grafted mesencephalic DA neuron survival. Unilaterally lesioned male Fischer 344 rats were grafted with either a suspension of mesencephalic cells or with both mesencephalic and striatal cell suspensions. Co-grafts were either mixed together or placed separately into the striatum. Lesioned rats receiving no graft served as controls. Rotational behavior was assessed following amphetamine challenge at 2 weeks prior to grafting and at 4 and 8 weeks following grafting. Only rats receiving co-grafts of nigral and striatal suspensions separated by a distance of 1 mm showed significant behavioral recovery from baseline rotational asymmetry. Both mixed and separate striatal co-grafts were associated with a doubling of DA neuron survival compared with solo mesencephalic grafts. In the mixed co-graft experiment, DA neurite branching appeared enhanced and TH-rich patches were observed, whereas with co-grafts that were separated, TH+ innervation of the intervening host striatum was increased significantly. These results provide the first evidence suggesting that nigral-striatal co-grafts, particularly those placed separately and in proximity to each other, increase both DA neuron survival and neurite extension from the mesencephalic component of the grafts.

Improvements in MPTP-induced deficits were only found in subjects that received fetal substantia nigra transplants into the caudate nucleus. The MPTP-induced deficits were assessed using an object retrieval task that examined cognitive and subtle motor performance and by behavioral observation to determine the overall status of the subjects. Subjects that were also moderately or severely impaired by MPTP administration but that received inappropriate donor cells or implant sites (cerebellum to CN or SN to cortex) did not show any evidence of behavioral recovery. These subjects could not respond on the task in the months after grafting and were sacrificed, showing no improvements in parkinsonian signs or healthy behavior signs, up to 5-6 months after surgery. Grafting of SN cells into the striatum of non-MPTP lesioned subjects failed to modify normal behavior or induce abnormal behavior determined by our 2 behavioral assessment methods. In those monkeys that received the appropriate transplants, TH immunohistochemistry revealed that cells of the fetal substantia nigra grafted into the caudate nucleus survived and extended neurites into the host striatum. Indeed, grafted dopamine neurons were often associated with appreciable innervation of the caudate nucleus and appeared to be well incorporated into the host brain. In contrast, examination of the striatum of subjects in the inappropriate-graft group (e.g., cerebellar cells grafted into the caudate) showed no evidence of TH staining within the graft or host caudate nucleus. This indicated that there was no evidence of dopamine neurons present in the grafted tissue and that the mere presence of a fetal graft did not appear to induce sprouting in these MPTP-treated subjects. Although behavioral recovery occurred in only those monkeys that received appropriate transplants (fetal SN to host CN) and not in those that received inappropriate grafts (fetal cerebellum to CN or fetal SN to cortex), the CSF HVA levels did not distinguish those monkeys with improved parkinsonism from those that remained severely parkinsonian. The finding that in some SN-CN grafted subjects reported here, there was evidence of increased dopamine and lowered HVA/dopamine ratio in the vicinity of the SN grafts (cf. Elsworth et al., 1990b) is consistent with the hypothesis that graft-derived or graft-induced dopamine production is responsible for behavioral recovery. In addition, the finding that CSF HVA levels in non-MPTP lesioned subjects were unchanged by fetal SN grafts further indicates that CSF HVA levels may not be sufficiently sensitive to changes in central dopamine production to reflect release of dopamine from relatively small grafts that may, in lesioned subjects, modify behavior.(ABSTRACT TRUNCATED AT 400 WORDS)

Evidence is presented indicating that norepinephrine (NE) inhibits vasopressin (VP) release from the rat hypothalamo-neurohypophyseal explant under some, but not all, conditions in vitro. NE at 10(-5) M inhibited basal VP release and inhibited acetylcholine-induced release in a concentration-dependent fashion. However, the induction of VP release caused by the addition of NaCl (sufficient to yield a 10 mosmol/kg H2O increase in culture medium osmolality) was not reduced by NE in concentrations as high as 10(-5) M. An alpha-adrenergic receptor mediation of the inhibition of VP release by NE was suggested by the ability of phentolamine and phenoxybenzamine, but not propranolol, to block this effect. In addition, phentolamine at 10(-4) M, but not equimolar amounts of propranolol, increased VP release when added alone on day 2 of culture, but not on days 3 or 4. Histofluorescence examination of additional explants revealed that endogenous catecholamine was still present on day 2 within varicosities in the supraoptic nucleus, but diminished by days 3 and 4, suggesting that endogenous NE could influence basal VP release. The results indicate that NE can inhibit spontaneous and cholinergically stimulated VP release from the hypothalamo-neurohypophyseal explant, but osmotic stimulation renders the explants insensitive to attenuation of VP release by NE.

The effect of the monosodium glutamate (MSG) induced lesion of the arcuate nucleus on catecholamines in the arcuate nucleus and median eminence of the mouse hypothalamus was determined using the Falck-Hillarp histofluorescence technique. The number of fluorescent perikarya in the arcuate nucleus of treated animals was decreased approximately 60%; the fluorescence intensity of surviving neurons was notably reduced. These changes were accompanied by a reduction in the intensity of fluorescence in the median eminence. Pretreatment of control and MSG-lesioned animals with a monoamine oxidase inhibitor (pargyline) greatly increased fluroescence in the median eminence and arcuate nucleus of both groups. However, the number of fluorescing perikarya of the arcuate nucleus of the normal pargyline treated group far exceeded that of the pargyline MSG animals. It is concluded that neonatally administered MSG caused destruction of a large number of dopaminergic arcuate perikarya.

A new method for the simultaneous visualization of brain peptides and monoamine neurotransmitters was employed to analyze the integrated morphology of hypothalamic catecholamines and neurophysins in young and old monkeys. Immunocytochemical analysis using bovine neurophysin revealed a dual papulation of light and dark stained cells in the paraventricular nucleus in young and old macaques. In general, both populations of neurons stained with less density in old macaques indicating the possibility of a reduced content of neurophysin. Further analysis using specific neurophysin antisera for vasopressin or oxytocin revealed an appreciable decrease in the number of vassopressin-containing perikarya in the 20 year monkey whereas oxytocin-synthesizing neurons did not show a similar change in numbers with age. Qualitatively, terminal innervation patterns of hypothalamic catecholamines remained strikingly constant in spite of marked reductions in dark-stained neurophysin perikarya of the paraventricular nucleus.

C57Bl/6N male mice at 4, 12, 20, and 28 months of age were processed for formaldehyde-induced fluorescence, and tuberoinfundibular dopamine (A-12) neurons were examined for qualitative, age-related changes. A-12 perikarya were weakly fluorescent at all ages studied. In mice of 4 and 12 months of age, A-12 terminals formed a brightly fluorescent band that filled the external zone of the median eminence. The zone of A-12 terminal fluorescence within the median eminence was less extensive and reduced in fluorescence intensity in 20-month old mice; in 28-month old mice, only a thin rim of fluorescence remained at the ventral edge of the median eminence. The drop in fluorescence intensity of A-12 terminals within the median eminence of the aged C57Bl/6N male mouse paralleled the decrease in A-12 terminal fluorescence previously reported for the Fischer 344 male rat [26]; however, mouse A-12 perikarya did not exhibit the dramatic increase in fluorescence intensity seen in the rat. The possibility that the divergent aging patterns exhibited in A-12 neurons of these two species might be related to differential resilience of their reproductive axes is discussed.

The relative content of monoamines within identified brain stem neurons in 4 and 20-year old monkeys (Macaca nemestrina) was determined with microspectrofluorometric techniques. Intraneuronal monoamine content was found to decrease with age in the locus coeruleus, substantia nigra and raphe and was found to be reduced further in the locus coeruleus by the presence of lipofuscin granules. These data indicate that intraneuronal monoamine content is reduced in the aged macaque.

Vasopressin (VP)-containing neurons of the supraoptic and paraventricular nuclei are influenced by a number of diverse, chemically defined neural systems. Of these the anatomy and physiology of noradrenergic and cholinergic afferents are best understood. Norepinephrine (NE)-containing neurons of the caudal medulla heavily innervate VP neurons; NE has been shown to either inhibit or facilitate the release of VP under different experimental paradigms. Acetylcholine is a powerful stimulant of VP release. gamma-Aminobutyric acid also appears to inhibit release. Several other putative transmitters are localized within the supraoptic and paraventricular nuclei, which suggests that VP release is integrated by a wide variety of neural systems.