6-hydroxydopa (6-OHDOPA) was synthesized with the expectation that it would be able to cross the blood-brain barrier to be enzymatically decarboxylated to 6-hydroxydopamine (6-OHDA), the newly discovered neurotoxin for noradrenergic and dopaminergic neurons. In part, 6-OHDOPA fulfilled these criteria. When administered experimentally to rodents, 6-OHDOPA destroyed peripheral sympathetic noradrenergic nerves and did exert neurotoxicity to noradrenergic nerves in brain—in large part, from its conversion to 6-OHDA. However, the efficacy of 6-OHDOPA was less than that of 6-OHDA; also, 6-OHDOPA was relatively selective for noradrenergic neurons; near-lethal doses of 6-OHDOPA were required to damage dopaminergic nerves; and ultimately, 6-OHDOPA was found to be an agonist at AMPA receptors, thus accounting for more non-specificity. Nevertheless, 6-OHDOPA was found to be a particularly valuable tool in uncovering processes and mechanisms associated with noradrenergic nerve regeneration and sprouting, particularly when administered to perinatal rodents. Also, 6-OHDOPA was a good tool for selective mapping of noradrenergic nerve tracts in brain, since dopaminergic tracts were unaffected and did not interfere with the histofluorescent methodology used for this purpose in the early 1970s. As an experimental research tool, 6-OHDOPA was valuable in a short time-window, but its utility is largely limited because of newer research technologies that provide better means today for nerve tract mapping, and for experimental approaches engaged toward study of processes and mechanisms attending nerve regeneration. AMPA actions of 6-OHDOPA have not been extensively studied, so this avenue may enliven use of 6-OHDOPA in the future.

The neonatally 6-hydroxydopamine (n6-OHDA)-lesioned rat has been the standard for 40 years, as an animal model of attention-deficit hyperactivity disorder (ADHD). Rats so lesioned during postnatal ontogeny are characterized by ∽99% destruction of dopaminergic nerves in pars compacta substantia nigra, with comparable destruction of the nigrostriatal tract and lifelong ∽99 % dopaminergic denervation of striatum, with lesser destructive effect on the ventral tegmental nucleus and associated lesser dopaminergic denervation of nucleus accumbens and prefrontal cortex. As a consequence of striatal dopaminergic denervation, reactive serotoninergic hyperinnervation of striatum ensues. The striatal extraneuronal milieu of DA and serotonin is markedly altered. Also, a variety of sensitization changes occur for dopaminergic D1 and D2 receptors, and for serotoninergic receptors. Behaviorally, these rats in adulthood display spontaneous hyperlocomotor activity, attentional deficits, and cognitive impairment-all of which are acutely attenuated by the psychostimulants amphetamine (AMPH) and methylphenidate (MPH) (i.e.,opposite to the acute effects of AMPH and MPH in intact control rats). The acute behavioral effects of AMPH and MPH in intact and lesioned rats are analogous to their respective acute effects in non-ADHD and in ADHD humans. The neurochemical template of brain, and behavioral series of changes in n6-OHDA-lesioned rats, is described in the review. Despite the fact that nigrostriatal damage is not an underlying pathophysiological process of human ADHD (i.e.,lacking construct validity), the described animal model has face validity (behavioral profile) and predictive validity (mirror of ADHD/MPH effects, as well as putative and new ADHD treatment effects). Also described in this review is a modification of the n6-OHDA rat, produced by adulthood partial lesioning of the serotoninergic fiber overgrowth. This ADHD model has even more accentuated hyperlocomotor and attentional deficits, counteracted by AMPH-thus providing a more robust means of animal modeling of ADHD. The n6-OHDA rat as a model of ADHD continues to be important in the search for new ADHD treatments.

The classic rodent model of Parkinson’s disease (PD) is produced by unilateral lesioning of pars compacta substantia nigra (SNpc) in adult rats, producing unilateral motor deficits which can be assessed by dopamine (DA) D2 receptor (D2-R) agonist induction of measurable unilateral rotations. Bilateral SNpc lesions in adult rats produce life-threatening aphagia, adipsia, and severe motor disability resembling paralysis-a PD model that is so compromised that it is seldom used. Described in this paper is a PD rodent model in which there is bilateral 99% loss of striatal dopaminergic innervation, produced by bilateral intracerebroventricular or intracisternal 6-hydroxydopamine (6-OHDA) administration to perinatal rats. This procedure produces no lethality and does not shorten the life span, while rat pups continue to suckle through the pre-weaning period; and eat without impairment post-weaning. There is no obvious motor deficit during or after weaning, except with special testing, so that parkinsonian rats are indistin-guishable from control and thus allow for behavioral assessments to be conducted in a blinded manner. L-DOPA (L-3,4-dihydroxyphenylalanine) treatment increases DA content in striatal tissue, also evokes a rise in extraneuronal (i.e.,in vivo microdialysate) DA, and is able to evoke dyskinesias. D2-R agonists produce effects similar to those of L-DOPA. In addition, effects of both D1-and D2-R agonist effects on overt or latent receptor supersensitization are amenable to study. Elevated basal levels of reactive oxygen species (ROS), namely hydroxyl radical, occurring in dopaminergic denervated striatum are suppressed by L-DOPA treatment. Striatal serotoninergic hyperinnervation ensuing after perinatal dopaminergic denervation does not appear to interfere with assessments of the dopaminergic system by L-DOPA or D1-or D2-R agonist challenge. Partial lesioning of serotonin fibers with a selective neurotoxin either at birth or in adulthood is able to eliminate sero-toninergic hyperinnervation and restore the normal level of serotoninergic innervation. Of all the animal models of PD, that produced by perinatal 6-OHDA lesioning provides the most pronounced destruction of nigrostriatal neurons, thus representing a model of severe PD, as the neurochemical outcome resembles the status of severe PD in humans but without obvious motor deficits.

The molecular mechanism responsible for degenerative process in the nigrostriatal dopaminergic system in Parkinson’s disease (PD) remains unknown. One major advance in this field has been the discovery of several genes associated to familial PD, including alpha synuclein, parkin, LRRK2, etc., thereby providing important insight toward basic research approaches. There is an consensus in neurodegenerative research that mitochondria dysfunction, protein degradation dysfunction, aggregation of alpha synuclein to neurotoxic oligomers, oxidative and endoplasmic reticulum stress, and neuroinflammation are involved in degeneration of the neuromelanin-containing dopaminergic neurons that are lost in the disease. An update of the mechanisms relating to neurotoxins that are used to produce preclinical models of Parkinson´s disease is presented. 6-Hydroxydopamine, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, and rotenone have been the most wisely used neurotoxins to delve into mechanisms involved in the loss of dopaminergic neurons containing neuromelanin. Neurotoxins generated from dopamine oxidation during neuromelanin formation are likewise reviewed, as this pathway replicates neurotoxin-induced cellular oxidative stress, inactivation of key proteins related to mitochondria and protein degradation dysfunction, and formation of neurotoxic aggregates of alpha synuclein. This survey of neurotoxin modeling—highlighting newer technologies and implicating a variety of processes and pathways related to mechanisms attending PD—is focused on research studies from 2012 to 2014.

Quinpirole-induced vertical jumping is a phenomenon first observed in rats treated from birth, once a day for 21 days or more, with the dopamine D 2 receptor agonist quinpirole. This quinpirole-induced behavioral sensitization is known as a priming process. To determine whether dopaminergic innervation influenced this priming phenomenon, groups of rats were lesioned at 3 days after birth with the neurotoxin 6-hydroxydopamine (6-OHDA; 67 μg in each lateral ventricle; desipramine pretreatment, 20 mg/kg ip, 1 h). Rats were additionally treated daily from birth with quinpirole HCl (3.0 mg/kg ip, salt form). Controls received saline vehicle in place of 6-OHDA and/or quinpirole. When rats were placed in individual observation cages (1 h acclimation) starting at 20 days after birth, acute quinpirole treatment produced vertical jumping in the quinpirole-primed group; and the effect persisted through the twenty-ninth day. In rats additionally lesioned with 6-OHDA, vertical jumping was enhanced at 20, 24, 26/27, and 28/29 day-with there being as much as a 32-fold increase in vertical jumping versus the group that was primed with quinpirole, but not lesioned with 6-OHDA. This finding indicates that an ontogenetic 6-OHDA lesion enhances quinpirole-induced vertical jumping in rats and that dopaminergic innervation may normally exert a suppressive effect on vertical jumping.

Rats lesioned shortly after birth with 6-hydroxydopamine (6-OHDA; 134 μg icv) represent a near-ideal model of severe Parkinson's disease because of the near-total destruction of nigrostriatal dopaminergic fibers. The element manganese, an essential cofactor for many enzymatic reactions, itself in toxic amount, replicates some clinical features similar to those of Parkinson's disease. The aim of this study was to examine the effect of neonatal manganese exposure on 6-OHDA modeling of Parkinson's disease in rats. Manganese (MnCl 2·4H 2O) 10,000 ppm was included in the drinking water of pregnant Wistar rats from the time of conception until the 21st day after delivery, the age when neonatal rats were weaned. Control rats consumed tap water. Other groups of neonatal rat pups, on the 3rd day after birth, were pretreated with desipramine (20 mg/kg ip 1 h) prior to bilateral icv administration of 6-OHDA (30, 60, or 137 μg) or its vehicle saline-ascorbic (0.1%) (control). At 2 months after birth, in rats lesioned with 30, 60, or 134 μg 6-OHDA, endogenous striatal dopamine (DA) content was reduced, respectively, by 66, 92, and 98% (HPLC/ED), while co-exposure of these groups to perinatal manganese did not magnify the DA depletion. However, there was prominent enhancement of DA D 1 agonist (i.e., SKF 38393)-induced oral activity in the group of rats exposed perinatally to manganese and also treated neonatally with the 30 mg/kg dose of 6-OHDA. The 30 mg/kg 6-OHDA group, demonstrating cataleptogenic responses to SCH 23390 (0.5 mg/kg) and haloperidol (0.5 mg/kg ip), developed resistance if co-exposed to perinatal manganese. In the group exposed to manganese and lesioned with the 60 mg/kg dose of 6-OHDA, there was a reduction in D2 agonist (i.e., quinpirole, 0.1 mg/kg)-induced yawning. The series of findings demonstrate that ontogenetic exposure to manganese results in an enhancement of behavioral toxicity to a moderate dose of 6-OHDA, despite the fact that there is no enhanced depletion of striatal DA depletion by the manganese treatment.

In the 1950s, the discovery of anti-nerve growth factor, an immunotoxin stunting sympathetic neural development, signaled the advent of neurotoxins as research modalities. Other selective neurotoxins were discovered in rapid succession. In the 1960s, 6-hydroxydopamine and 6-hydroxydopa were shown to destroy noradrenergic and dopaminergic nerves. Excitotoxins (glutamate, aspartate, and analogs) were discovered in the 1970s. DSP-4 [N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine] proved to be selective for noradrenergic destruction, while 5,6-and 5, 7-dihydroxytryptamines were relatively selective for serotonin neurons. Additional neurotoxins were discovered, but it was MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) that predominated neurotoxicity research in the 1980s. Eventually, Clostridium botulinum neurotoxin (BoNT), discovered as a "poisonous" principle in the late 1800s, resurfaced in purified and standardized forms as a clinically useful drug. Neurotoxins represent chemical tools, useful not only for discerning neuronal mechanisms and animal modeling of neurological disorders, but also for their use in medicine and potential as treatments for medical disorders. This unit reviews the early discovery of neurotoxins, describes categories of neurotoxins, and finally characterizes their usefulness - first as research tools, and eventually as clinical therapeutic agents.

Although it is well-recognized that catecholamines are generally unable to penetrate the developed blood-brain barrier (BBB) to gain entry into brain, except at circumventricular sites where the BBB is absent or deficient, onto-genetic development of this barrier seems to have escaped systematic study. To explore BBB development, several approaches were used. In the first study rats were treated once on a specific day of postnatal ontogeny, as early as the day of birth, with the neurotoxin 6-hydroxydopa-mine (6-OHDA; 60 mg/kg), and then terminated in adulthood for regional analysis of endogenous norepinephrine (NE) content of brain. In another study, rats were treated once, on a specific day of postnatal ontogeny, with the BBB-perme-able neurotoxin 6-hydroxydopa (6-OHDOPA; 60 mg/kg) following pretreatment with the BBB-impermeable amino acid decarboxylase inhibitor carbidopa (100 mg/kg IP), then terminated in adulthood for regional analysis of endogenous NE content of brain. In the third study rats were treated once, on a specific day of postnatal ontogeny, with the analog [3H]metaraminol, and terminated 1 hour later for determination of regional distribution of tritium in brain. On the basis of [3H]metaraminol distribution and NE depletions after neurotoxin treatments, it is evident that the BBB in neocortex, striatum, cerebellum and other brain regions forms in stages over a period of at least 2 weeks from birth. Moreover, because the BBB consists of several element (physical-, ion-restrictive-, and enzymatic-barrier), the method employed will derive data mainly applicable to the targeted aspect of the barrier, which may or may not necessarily coincide with elements of the barrier that have a different rate of ontogenetic development. Accordingly, it is evident that some aspects of physical- and ion-restrictive elements of the BBB form within approximately the first week after birth in rat neocortex and striatum, while enzymatic elements of the BBB form more than than 2 week later. Regardless, the BBB forms at earlier times in forebrain vs hindbrain regions.

In the neonatally 6-hydroxydopamine (6-OHDA)-lesioned rat hyperlocomotor activity, first described in the 1970s, was subsequently found to be increased by an additional lesion with 5,7-dihydroxytryptamine (5,7-DHT) (i.c.v.) in adulthood. The latter animal model (i.e., 134 μg 6-OHDA at 3 d postbirth plus 75 μg 5,7-DHT at 10 weeks; desipramine pretreatments) was used in this study, in an attempt to attribute hyperlocomotor attenuation by D,L-amphet-amine sulfate (AMPH) and m-chlorophenylpi-perazine di HCl (mCPP), to specific changes in extraneuronal (i.e., in vivo microdialysate) levels of dopamine (DA) and/or serotonin (5-HT). Despite the 98-99% reduction in striatal tissue content of DA, the baseline striatal microdialysate level of DA was reduced by 50% or less at 14 weeks, versus the intact control group. When challenged with AMPH (0.5 mg/kg), the microdialysate level of DA went either unchanged or was slightly reduced over the next 180 min (i.e., 20 min sampling), while in the vehicle group and 5,7-DHT (alone) lesioned group, the microdialysate level was maximally elevated by ∼225% and ∼450%, respectively - and over a span of nearly 2 h. Acute challenge with mCPP (1 mg/kg salt form) had little effect on microdialysate levels of DA, DOPAC and 5-HT. Moreover, there was no consistent change in the microdialysate levels of DA, DOPAC, and 5-HT between intact, 5-HT-lesioned rats, and DA-lesioned rats which might reasonably account for an attenuation of hyperlocomotor activity. These findings indicate that there are other important neurochemical changes produced by AMPH-and mCPP-attenuated hyperlocomotor activity, or perhaps a different brain region or multiple brain regional effects are involved in AMPH and mCPP behavioral actions.

To determine whether behavioral sensitization produced by prolonged D-amphetamine administration affects susceptibility of nigrostriatal dopaminergic neurons to the neurotoxic actions of 6-hydroxydopamine (6-OHDA), rats were treated daily from the 23 rd day after birth for 11 consecutive days with D-amphetamine (1.0 mg/kg s.c.) or saline. On the last day of treatment, one group primed with D-amphetamine and one control group of rats were tested to confirm behavioral sensitization development. The remaining animals were additionally treated on the 34 th day (one day after the last D-amphetamine injection) with 6-OHDA HBr (300 μg in 10 μl i.c.v., salt form, half in each lateral ventricle) or its vehicle. Four weeks later the levels of dopamine (DA) and its metabolites 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 3-metoxytyramine (3-MT), as well as 5-hydroxytrypatmine (5-HT) and its metabolite 5-hydroxyindoleacteic acid (5-HIAA) were assayed in the striatum, by HPLC/ED. In rats with behavioral sensitization, 6-OHDA reduced endogenous dopamine and its metabolites content to a comparable degree in comparison to controls. This finding indicates that presumed up-regulation of the dopamine transporter in the behaviorially sensitized rats did not increase the neurotoxicity of a high dose of 6-OHDA.