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Studies of dopamine (DA) clearance and diffusion were carried out after unilateral destruction of striatal DA afferents using intraparenchymal injection of 6-hydroxydopamine (6-OHDA). Rats were screened for the extent of DA depletion by testing for contralateral rotations induced by 0.05 mg/kg of apomorphine. Exogenous DA clearance and diffusion were determined using rapid (5 Hz) in vivo chronoamperometry with Nafion-coated carbon-fiber electrodes. Local applications of DA by pressure ejection from micropipettes into the lesioned vs. nonlesioned striata, with dose adjusted to elicit roughly equivalent amplitudes of electroactive signals, manifested a much prolonged clearance time on the lesioned side. A second protocol involved administration of equal volumes (amounts) of DA into lesioned vs. nonlesioned striata. In such cases, a volume of DA which manifested no detectable signal on the intact side produced a pronounced electrochemical signal on the lesioned side. When nomifensine, a high-affinity DA uptake inhibitor, was locally applied into the intact striatum, a subsequent ejection of DA produced a much larger signal than when the same volume was given before nomifensine. Very little or no effects of nomifensine were seen in 6-OHDA-lesioned striata. Taken together, these data indicate a much prolonged clearance time of DA in 6-OHDA-denervated striata. Moreover, the data also suggest that the primary mechanism underlying this effect is the loss of high affinity neuronal uptake. Thus, such changes in DA clearance may help account for some of the well documented compensatory phenomena which occur after DA-depleting lesions in animals and in man.

The rat beta nerve growth factor (NGF) gene was inserted into a mammalian expression vector and cotransfected with a plasmid conferring resistance to neomycin into mouse 3T3 fibroblasts. From this transfection a stable cell line was selected that contains several hundred copies of the rat NGF gene and produces excess levels of recombinant NGF. Such genetically modified cells were implanted into the rat brain as a probe for in vivo effects of NGF on central nervous system neurons. In a model of the cortical cholinergic deficits in Alzheimer disease, we demonstrate a marked increase in the survival of, and fiber outgrowth from, grafts of fetal basal forebrain cholinergic neurons, as well as stimulation of fiber formation by intact adult intrinsic cholinergic circuits in the cerebral cortex. Adult cholinergic interneurons in intact striatum also sprout vigorously toward implanted fibroblasts. Our results suggest that this model has implications for future treatment of neurodegenerative diseases.

The presence of nerve growth factor (NGF) mRNA and protein in the rat central nervous system is documented. Blot-hybridization analysis showed an abundance of NGF mRNA in the hippocampus, cerebral cortex, and olfactory bulb. Enzyme immunoassay confirmed significant levels of a NGF-like protein in the hippocampus and cerebral cortex. Bioassay of a NGF-like immunoaffinity-purified protein from these regions was physiologically indistinguishable from NGF. Immunohistochemistry revealed a widespread distribution of NGF-like reactivity in the adult brain, preferentially in fiber tracts. NGF mRNA accumulation began at birth, with adult levels reached 3 weeks postnatally. Enzyme immunoassay detected the presence of a NGF-like protein in the embryonic rat brain. Postnatally, the level of NGF-like protein reached a maximum at 3 weeks. Additionally, a distinct fetal form of NGF may exist.

The long-term (i.e., 4-5 months) effects of large doses (3 X 50 mg/kg) of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on striatal dopamine-containing afferents were studied in the NMRI strain of mice. Recently improved in vivo electrochemical methods were first used to examine the magnitude, spatial distribution and temporal dynamics of monoamine release initiated via local application of potassium in various regions of the mouse striatum. Immunohistochemical localization of tyrosine hydroxylase and computer-based image analysis were also used to quantitate regional catecholamine-containing nerve fiber densities in the caudate nucleus. The in vivo electrochemical studies showed a statistically significant decrease in the average potassium-evoked release of electroactive species from the MPTP-treated mouse caudate nucleus vs. control. Greater decreases in release were seen in dorsal than in ventral striatum (55% vs. 33%). The average rise time of potassium-evoked release was also significantly prolonged (greater than 50%) after MPTP pretreatment. Histochemical studies showed an overall reduction in the density of dopamine-containing terminals in the drug-treated mice, with a greater loss observed in the more dorsal regions of the caudate nucleus. The experimental data thus support a long-term selective destruction of dorsal vs. ventral dopamine-containing afferents to the striatum by the neurotoxin MPTP in mice.

Human cortex cerebri and cerebelli xenografts from first-trimester fetal tissue fragments were used to study the effects of ethanol on single human central neurons. Transplants were placed into the anterior eye chamber of athymic nude rats and allowed to develop for 3 to 11 months. Immunohistologic analysis revealed graft structures that stained positively for a number of neuronal, transmitter-related, glial and vascular markers. Superfusion of ethanol (EtOH) elicited a reversible and dose-dependent depression of action potential discharge. At least two populations of neurons could be identified--a more sensitive group with an EC50 of 3.0 mM and a less sensitive group with an EC50 of 22.4 mM. These EtOH levels are within the range eliciting behavioral signs of intoxication in humans. EtOH-induced depressions could be antagonized by administration of the benzodiazepine inverse agonist Ro 15-4513. This study represents the first demonstration, to our knowledge, of the electrophysiologic actions of EtOH on single neurons from human brain, and provides dose-response data collected with known concentrations of EtOH as well as evidence for the blockade of these EtOH effects by the Roche compound.