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Background: To determine if the intranasal delivery of neuroactive compounds is a viable, long-term treatment strategy for progressive, chronic neurodegenerative disorders, such as Parkinson's disease (PD), intranasal methodologies in preclinical models comparable to humans are needed., New Method: We developed a methodology to evaluate the repeated intranasal delivery of neuroactive compounds on the non-human primate (NHP) brain, without the need for sedation. We evaluated the effects of the neuroactive peptide, DNSP-11 following repeated intranasal delivery and dose-escalation over the course of 10-weeks in Rhesus macaques. This approach allowed us to examine striatal target engagement, safety and tolerability, and brain distribution following a single 125 I-labeled DNSP-11 dose., Results: Our initial data support that repeated intranasal delivery and dose-escalation of DNSP-11 resulted in bilateral, striatal target engagement based on neurochemical changes in dopamine (DA) metabolites-without observable, adverse behavioral effects or weight loss in NHPs. Furthermore, a 125 I-labeled DNSP-11 study illustrates diffuse rostral to caudal distribution in the brain including the striatum-our target region of interest., Comparison With Existing Methods: The results of this study are compared to our experiments in normal and 6-OHDA lesioned rats, where DNSP-11 was repeatedly delivered intranasally using a micropipette with animals under light sedation., Conclusions: The results from this proof-of-concept study support the utility of our repeated intranasal dosing methodology in awake Rhesus macaques, to evaluate the effects of neuroactive compounds on the NHP brain. Additionally, results indicate that DNSP-11 can be safely and effectively delivered intranasally in MPTP-treated NHPs, while engaging the DA system., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Glial cell line-derived neurotrophic factor (GDNF) promotes recovery of the injured nigrostriatal dopamine system and improves motor functions in both rodent and nonhuman primate models of Parkinson's disease (PD). The neurorestorative effects of a single administration of GDNF last for at least 1 month and can be maintained in rhesus monkeys by monthly injections. Adult midbrain dopamine neurons stimulated by GDNF show increased cell size, neurite extent, and expression of phenotypic markers. In parkinsonian nonhuman primates, GDNF treatment improves three of the cardinal features of PD: bradykinesia, rigidity, and postural instability. Although intracerebral administration is necessary because of the blood-brain barrier, intraventricular, intrastriatal, and intranigral routes of administration have been found to be efficacious in rodents and nonhuman primates. GDNF also induces neuroprotective changes in dopamine neurons which are active within hours after trophic factor administration. The powerful neuroprotective and neurorestorative properties of GDNF seen in preclinical studies suggest that trophic factors may play an important role in treating PD.

The Brown Norway/Fischer 344 F1 hybrid rats (F344BNF1) is a newer rat model and is emerging as an important rodent model of aging. In the present study we used motoric performance tests, intracerebral microdialysis, and neurochemical measures of postmortem brain tissue to investigate the effects of aging in young (4-5 months), middle-aged (18-19), and old (24-25 months) F344BNF1 hybrid rats. We observed that old F344BNF1 rats exhibited decreased motoric performance, and lower levels of spontaneous and d-amphetamine-induced locomotor activity than those observed in young F344BNF1 rats. Microdialysis measures of extracellular basal levels of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), and 4-hydroxy-3-methoxyphenylacetic acid (HVA) were significantly diminished in the striata of the middle-aged and old rats as compared to levels in young animals. In addition, d-amphetamine-evoked overflow of DA was significantly decreased in the middle-aged and aged rat striatum as compared to DA overflow in young F344BNF1 rats. Studies of postmortem brain tissue showed that the changes in overflow of DA correlated with significantly lower DA tissue content in ventral striatum and midbrain. Moreover, both dopamine turnover ratios (DOPAC/DA, HVA/DA) and the serotonin turnover ratio (5-HIAA/5-HT) were significantly elevated in the ventral striatum and nucleus accumbens. The results of this study demonstrate a correlation between reductions in striatal DA neurochemistry and diminished motor function in aged F344BNF1 rats., (Copyright 1998 Elsevier Science B.V.)

Adrenal medulla has often been used as a donor tissue for transplantation into damaged central nervous system, with functional effects ranging from very good to nonexistent. The grafts have often been associated with morphological evidence of stimulated recipient dopaminergic fiber plasticity. The interpretation of these results has been difficult due to variable but mostly poor graft survival. The present study combines two experiments which evaluated the effects of intrastriatal xenogeneic adrenal medullary cell suspension grafts on rat recipients. First, bovine adrenal medulla cell suspension grafts of various compositions were tested for their functional and morphologic effects on immunosuppressed hemiparkinsonian rats. In the second experiment, graft rejection was allowed to occur in half of the rats in order to determine a possible contribution of the inflammatory/immune response to increased dopaminergic fiber plasticity of the recipient. At 28 days, grafts of all cell types survived well in immunosuppressed rats, but none of the grafted cell types was associated with either an amelioration of amphetamine-induced rotation or an increase in striatal tyrosine hydroxylase immunoreactivity around the graft site. The latter phenomenon was observed only in the nonimmunosuppressed rats with rejected grafts. Our findings strongly support the role of inflammatory/immune response to grafting in stimulating dopaminergic fiber plasticity and in the appearance of sprouting.

Rapid (5 Hz) chronoamperometric recordings using Nafion-coated carbon fiber electrodes (30-90 microns o.d.) combined with pressure-ejection of potassium from micropipettes were used to investigate potassium-evoked overflow of dopamine (DA) in the striatum of young (5 to 10 years old) and middle-aged (19 to 23 years old) anesthetized rhesus monkeys. The potassium-evoked DA-like signals from the 19- to 23-year-old animals were significantly lower in amplitude than those recorded in the young animals. In addition, the temporal dynamics of DA signals in the caudate nucleus of middle-aged animals were faster, while the time courses of the signals recorded in the putamen of middle-aged monkeys were significantly longer as compared to the signals recorded from young animals. Moreover, home cage activity levels of the middle-aged animals were significantly lower. Taken together, these data support age-related changes in the output of DA from DA fibers in the striatum of middle-aged monkeys.

Injection of quinolinic acid in the rat striatum mimics neurochemical changes observed in Huntington's disease. We previously demonstrated that intrastriatal transplantation of fetal striatum or gelfoam protects against toxicity induced by a subsequent intrastriatal injection of quinolinic acid performed one week later. Herein, we examined whether fetal striatum or sham transplantation provides protection against quinolinic acid that lasts up to four weeks. Intrastriatal quinolinic acid injection produces neuronal loss and gliosis in Nissl staining, loss of cytochrome oxidase histochemical staining, decrease in autoradiographic binding of [3H]SCH 23390-labeled dopamine D1 and [3H]CGS 21680-labeled adenosine A2 receptors, and increase in autoradiographic binding of [3H]PK 11195-labeled peripheral benzodiazepine binding sites. None of these changes was observed in rats transplanted with fetal striatum one, two or four weeks before quinolinic acid injection. In animals transplanted with fetal striatal tissue, Nissl staining showed healthy grafts located in normal appearing striata. Although sham transplantation performed one week before quinolinic acid injection also protected against histological, histochemical and binding changes, sham transplantation performed two or four weeks before quinolinic acid injection was less effective in attenuating quinolinic acid-induced striatal toxicity. Thus, sham transplantation provides transient protection against quinolinic acid-induced striatal toxicity, whereas implantation of tissue such as fetal striatum seems to be required for long-lasting protection. Our study suggests that intracerebral transplantation may also act through other mechanisms than restoration of deficient neurotransmitters or damaged pathways, a finding which may have significant clinical implications in assessing the potential benefit of this approach for the treatment of neurodegenerative disorders such as Huntington's disease.

In vivo electrochemistry was used to examine the clearance of locally applied dopamine in the caudate nuclei of normal, control monkeys and in the non-lesioned and lesioned caudate nuclei of unilateral 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys. Chronoamperometric recordings were continuously made using Nafion-coated carbon fiber electrodes in anesthetized animals while a calibrated amount of dopamine was pressure ejected from a micropipette adjacent (250-300 microns) to the electrode. The dopamine signals recorded from the MPTP-lesioned caudate nuclei had a greater amplitude and time course than those recorded from both the controls and from the non-lesioned side of the treated animals, indicating a loss of high-affinity uptake in the lesioned caudate. However, the time course was faster, and the clearance rate greater, in signals recorded from the contralateral side of the treated animals when compared to control caudate nuclei. This suggests that there is an up-regulation in high-affinity dopamine uptake in the non-lesioned side of the MPTP-treated animals. This change may represent a compensatory mechanism that is attempting to maintain a balance in extracellular dopamine levels between the lesioned and non-lesioned sides.

Previous studies in our laboratory have shown that intrastriatal implants of fetal striatum significantly attenuate excitotoxic damage resulting from a 240 nmol quinolinic acid (QA) challenge delivered 7 days later. In contrast, animals with intrastriatal implants of other tissue types (adipose tissue, peripheral nerve or adrenal medulla) demonstrate a more limited, but consistent trend in protection from QA excitotoxicity. The present study was designed to test the hypothesis that partial striatal protection found in animals receiving peripheral tissue grafts is due to the transplantation procedure eliciting a host response which attenuates excitotoxicity. Adult female Long-Evans rats received either cellular (fetal striatum) or acellular (gelfoam) implants followed 1 week later by a unilateral injection of 240 nmol QA into the grafted striatum. Animals were tested for rotational asymmetries before grafting, post-implantation, and after lesioning. Compared to baseline rotational behavior, rats which received implants did not show changes in ipsilateral turning after QA lesions. This protective effect was not limited to rotational behavior since improvements in spontaneous locomotor activity were evident. In addition, the adipsia and aphagia often associated with striatal lesions were ameliorated in both groups of grafted animals. Morphometric analysis demonstrated that endogenous dopaminergic, cholinergic and enkephalinergic systems in the two transplanted groups sustained less excitotoxic damage than in the QA lesioned, non-grafted animals. These results are consistent with the hypothesis that a host generated response activated by the implantation procedure provides a protective effect against QA excitotoxicity.

Administration of the neurotoxin 6-hydroxydopamine (6-OHDA) to rat brain causes biochemical and neuroanatomical changes to the nigrostriatal dopaminergic pathway similar to those observed in Parkinson's disease (PD). Although the cause of PD is unknown, it has been hypothesized that the neurodegenerative changes seen in PD might result from exposure to a neurotoxin. Therefore, strategies for limiting neurotoxin-induced dopaminergic damages, like those caused by 6-OHDA, may be of both clinical and basic interest. Accordingly, we tested the ability of both fetal neural (striatum) and fetal non-neural (liver) tissue implants to protect the rat striatum against the toxic effects of a subsequent intrastriatal injection of 6-OHDA. Non-grafted rats (lesion only) showed amphetamine-induced rotational behavior and a decrease in striatal [3H]mazindol-labeled dopamine uptake sites after 6-OHDA injection. In contrast, the animals grafted with striatum or liver showed no behavioral or biochemical changes. Interestingly, sham-transplanted control animals were also protected against the 6-OHDA-induced toxicity. These results suggest that the resistance of the dopaminergic system against 6-OHDA neurotoxicity observed in grafted and sham-transplanted animals is likely to be related to the surgical procedure itself. This observation points to a possible role for surgery-related events in the clinical improvement described in PD patients who underwent intracerebral transplantation.

Quinolinic acid (QA) and related excitotoxins produce a pattern of neuronal loss and neurochemical changes in the rat striatum similar to that of patients suffering from Huntington's disease, suggesting neurotoxicity is important in the etiology of that disease. Thus, strategies for limiting excitotoxin-induced striatal damage, like that caused by QA, may be of great benefit to these individuals. Accordingly, we tested the ability of both neural and non-neural tissue implants to protect the rat striatum against a subsequent QA challenge. Our results demonstrated that recipients of fetal striatal grafts were significantly less affected by striatal injections of QA than non-grafted animals. In contrast to the latter, fetal striatal tissue recipients did not exhibit apomorphine-induced rotation behavior and showed a sparing of cholinergic and enkephalinergic systems normally lost following QA injections. Animals grafted with adult rat sciatic nerve, adrenal medulla or adipose tissue all showed a less dramatic behavioral protection and sparing of cholinergic and enkephalinergic systems. These results suggest that fetal striatal tissue exerts an optimal, and perhaps specific protective influence on the host brain.

The intrastriatal injection of the excitotoxin quinolinic acid (Q.A.) in rats produces neuroanatomical and neurochemical changes mimicking those appearing in the striatum in Huntington's disease (H.D.). Although its cause is unknown, it has been hypothesized that the neurodegenerative changes seen in H.D. may result from the action of an endogenous toxin. Therefore, the development of new strategies for limiting or preventing Q.A.-or other neurotoxic-induced degeneration may be of therapeutic interest for neurodegenerative disorders. Accordingly, we tested the ability of various tissue transplants to protect the rat striatum against a subsequent Q.A. insult. Using a "unilateral model", i.e. unilateral intrastriatal grafts followed by an ipsilateral intrastriatal injection of Q.A., we were able to quantify a behavioral protective effect of the grafts in recording the apomorphine-induced rotational behavior that normally appears after the striatal Q.A.-induced lesion. Our results show that one of the tested tissue, fetal striatum, protects the recipients against the lesioned-induced rotational behavior that appeared in non-grafted lesioned animals. The other grafted tissues (adrenal medulla, peripheral nerve, adipose tissue) seemed to provide a less dramatic protection than fetal striatum; however, this difference did not reach significance. Quantification of the striatal neuronal loss showed that the behavioral protection is significantly correlated with a better neuronal survival in the grafted animals. These results suggest that intracerebral grafts can protect the host brain against a toxin-induced damage, like the one resulting from Q.A. intrastriatal injection. Though fetal striatal grafts seem to exert an optimal protection, this protective effect may at least partially result from a host-mediated response to the transplantation procedure. The mechanism underlying this protective effect is unclear, but the present data suggest that it might be related to a transplantation-induced astroglial reaction resulting in an increased neuronotrophic activity that could protect against the toxic effect of Q.A. The results of this study also support the concept that the effect of transplantations could occur through processes other than a direct restoration of deficient transmitters or a reconstruction of damaged pathways. Further characterization of the factors implicated in the present paradigm might conceivably open avenues for possible therapeutic preventive interventions in neurodegenerative disorders.

Autopsy results on patients and corresponding studies in nonhuman primates have revealed that autografts of adrenal medulla into the striatum, used as a treatment for Parkinson's disease, do not survive well. Because adrenal chromaffin cell viability may be limited by the low levels of available nerve growth factor (NGF) in the striatum, the present study was conducted to determine if transected peripheral nerve segments could provide sufficient levels of NGF to enhance chromaffin cell survival in vitro and in vivo. Aged female rhesus monkeys, rendered hemiparkinsonian by the drug MPTP (n-methyl-4-phenyl-1,2,3,6 tetrahydropyridine), received autografts into the striatum using a stereotactic approach, of either sural nerve or adrenal medulla, or cografts of adrenal medulla and sural nerve (three animals in each group). Cell cultures were established from tissue not used in the grafts. Adrenal chromaffin cells either cocultured with sural nerve segments or exposed to exogenous NGF differentiated into a neuronal phenotype. Chromaffin cell survival, when cografted with sural nerve into the striatum, was enhanced four- to eightfold from between 8000 and 18,000 surviving cells in grafts of adrenal tissue only up to 67,000 surviving chromaffin cells in cografts. In grafts of adrenal tissue only, the implant site consisted of an inflammatory focus. Surviving chromaffin cells, which could be identified by both chromogranin A and tyrosine hydroxylase staining, retained their endocrine phenotype. Cografted chromaffin cells exhibited multipolar neuritic processes and numerous chromaffin granules, and were also immunoreactive for tyrosine hydroxylase and chromogranin A. Blood vessels within the graft were fenestrated, indicating that the blood-brain barrier was not intact. Additionally, cografted chromaffin cells were observed in a postsynaptic relationship with axon terminals from an undetermined but presumably a host origin.

Amitotic [3H]thymidine-labeled C6 glioma cells, which are known to produce neurotrophic factor(s), were grafted alone and with adrenal chromaffin cells in an attempt to improve chromaffin cell survival and phenotypic differentiation. Long-Evans rats with unilateral 6-hydroxydopamine-induced lesions of the nigrostriatal pathway were divided into four groups: (1) those receiving adrenal medullary cells co-transplanted with C6 glioma cells; (2) those receiving adrenal medullary graft alone; (3) those receiving C6 glioma grafts alone; and (4) those serving as a vehicle control group. All rats were killed one month after transplantation. Immunohistochemical, neurochemical, and autoradiographic methods were used to identify and characterize the grafted cells. Tyrosine hydroxylase-immunoreactive cells were found in all animals that received grafts of the adrenal medulla alone or of adrenal medulla co-transplanted with C6 glioma cells. The cograft recipients had more tyrosine hydroxylase-immunoreactive cells than the hosts receiving just adrenal chromaffin cells (P less than 0.05). Additionally, more grafted chromaffin cells formed processes in the former group. All three tissue recipient groups (adrenal medullary, C6 glioma cell, and cografted animals) had a significant reduction (P less than 0.05) in ipsilateral rotations after amphetamine (0.5 mg/kg i.p.) injections as compared to the control vehicle recipient group. Moreover, the reduction in rotation was more marked in the cografted hosts than in the other two implanted groups (P less than 0.05). Significantly higher dopamine levels were found in the transplant sites of both cograft and adrenal medullary graft recipients than in sham grafted control animals.

The present studies were undertaken to determine if MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) treatment in mice alters catecholamine or serotonin systems of the brain in addition to reported effects on the dopaminergic nigrostriatal system. Male Swiss-Webster mice were injected with 30 mg/kg MPTP daily for three days. Treated and control animals were sacrificed 10 and 24 days after the last injection, and brains were prepared for serotonin immunocytochemistry and catecholamine histofluorescence. MPTP treatment resulted in a reduced number of neurons and a reduced intensity of the fluorescence in the remaining cell bodies of substantia nigra pars compacta. A reduced presence of catecholamine varicosities in the medullary raphe nuclei and the ventromedial reticular formation of the medulla was observed. Immunocytochemical studies revealed no alteration in the number of serotonin positive cell bodies in the dorsal and median raphe nuclei. We suggest that MPTP treatment selectively alters some non-nigrostriatal catecholamine systems of the brain stem in addition to its toxic effects on the dopaminergic nigrostriatal system, while leaving other non-nigrostriatal catecholamine systems intact.

The drug, 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP), depletes striatal dopamine levels in primates and certain rodents, including mice, and produces parkinsonian-like symptoms in humans and nonhuman primates. To investigate the consequences of grafting adrenal medullary tissue into the brain of a rodent model of Parkinson's disease, a piece of adult mouse adrenal medulla was grafted unilaterally into mouse striatum 1 week after MPTP treatment. This MPTP treatment resulted in the virtual disappearance of tyrosine hydroxylase-immunoreactive fibers and severely depleted striatal dopamine levels. At 2, 4, and 6 weeks after grafting, dense tyrosine hydroxylase-immunoreactive fibers were observed in the grafted striatum, while only sparse fibers were seen in the contralateral striatum. In all cases, tyrosine hydroxylase-immunoreactive fibers appeared to be from the host rather than from the grafts, which survived poorly. These observations suggest that, in mice, adrenal medullary grafts exert a neurotrophic action in the host brain to enhance recovery of dopaminergic neurons. This effect may be relevant to the symptomatic recovery in Parkinson's disease patients who have received adrenal medullary grafts.

The survival and functional properties of dispersed cell implants of catecholaminergic cells obtained from the peripheral nervous system of adult rats (adrenal medulla and carotid body glomus cells) and PC12 cells from a rat pheochromocytoma cell line were examined following transplantation into the striatum of the adult rat. The host animals, all with unilateral 6-hydroxydopamine (6-OHDA) nigrostriatal lesions, were divided into 5 groups: (1) PC12 cells transplanted into Cyclosporin-A treated hosts; (2) PC12 cell grafts into hosts without Cyclosporin-A treatment; (3) grafts of adrenal medullary cells; (4) grafts of glomus cells; and (5) vehicle controls. All animals were sacrificed one month after transplantation. Immunocytochemical staining for tyrosine hydroxylase, the rate-limiting enzyme for catecholamine synthesis, was used to identify and characterize the grafted cells. PC12 cells were detected in four of six Cyclosporin-A treated rats, and two of these grafts developed into tumors. However, only one of the six non-Cyclosporin-A treated hosts was found to have surviving PC12 cells, and none of these rats developed tumors. No significant differences in rotational behavior were seen in either of the PC12 cell recipient groups. Grafted cells could be identified in all of the adrenal medullary and glomus cell recipients. However, the number of surviving cells was quite limited, with not more than 100 tyrosine hydroxylase-positive grafted cells found in any one recipient. Tyrosine hydroxylase-positive fibers were present adjacent to the transplants in these latter graft recipients, but the fibers appeared to be of host origin rather than from the grafts.(ABSTRACT TRUNCATED AT 250 WORDS)