Beneficial effects of n-3 polyunsaturated fatty acids administration in a partial lesion model of Parkinson's disease: The role of glia and NRf2 regulation.

Omega-3 polyunsaturated fatty acids (n-3 PUFAs) have been widely associated to beneficial effect over different neurodegenerative diseases. In the present study, we tested the potential therapeutic effect of docohexanoic acid (DHA) and its hydroxylated derivate, DHAH, in a partial lesion model of Parkinson's disease (PD). One month before and four months after the striatal lesion with 6-OHDA was made, the animals were daily treated with DHA (50 mg/kg), DHAH (50 mg/kg), vehicle or saline, by intragastric administration. Animal groups under n-3 PUFA treatments exhibited a trend to improve in amphetamine-induced rotations and cylinder test. The beneficial effect seen in behavioral studies were confirmed with TH immunostaining. TH+ fibers and TH+ neurons increased in the experimental groups treated with both n-3 PUFAs, DHA and DHAH. Moreover, the n-3 PUFAs administration decreased the astrogliosis and microgliosis, in both the striatum and substantia nigra (SN), with a higher decrease of GFAP+ and Iba-1+ cells for the DHAH treated group. This experimental group also revealed a positive effect on Nrf2 pathway regulation, decreasing the positive Nrf2 immmunostaining in the striatum and SN, which revealed a potential antioxidant effect of this compound. Taking together, these data suggest a positive effect of n-3 PUFAs administration, and more concretely of DHAH, for PD treatment as it exhibited positive results on dopaminergic system, neuroinflammation and oxidative stress.

Electroacupuncture Produces the Sustained Motor Improvement in 6-Hydroxydopamine-Lesioned Mice.

Clinical and research evidence has shown that electroacupuncture (EA) promotes recovery of motor function in patients with Parkinson's disease (PD). However, the "efficacy span" of EA treatment, especially the long-term effect of EA that is thought to last after the cessation of EA treatment, has not been investigated. The present study thus investigated and compared the effect of EA during and after chronic EA application on motor activity and dopamine lesions in a 6-hydroxydopamine (6-OHDA)-lesioned mouse model of PD. Chronic EA treatment (30 min a day, 6 days a week for 2 or 4 weeks) significantly attenuated motor deficiency and reduced dopamine neuron degeneration. Remarkably, EA showed a long-lasting effect after the cessation of EA stimulation. At 2 and 4 weeks after the termination of EA, EA continued to improve motor function in 6-OHDA-lesioned mice. Consistent with sustained behavioral effects, EA induced an enduring increase in the dopamine turnover ratio in the striatum 2 weeks after the cessation of EA treatment. Here we demonstrated that the therapeutic effect of EA outlasted the duration of EA application. During a relatively long period of time after the completion of EA treatment, EA is able to continue to improve motor function and enhance dopamine availability in 6-OHDA-lesioned PD mice.

Naringin treatment induces neuroprotective effects in a mouse model of Parkinson's disease in vivo, but not enough to restore the lesioned dopaminergic system.

We recently reported that treatment with naringin, a major flavonoid found in grapefruit and citrus fruits, attenuated neurodegeneration in a rat model of Parkinson's disease (PD) in vivo. In order to investigate whether its effects are universally applied to a different model of PD and whether its treatment induces restorative effects on the lesioned nigrostriatal dopaminergic (DA) projection, we observed the effects of pre-treatment or post-treatment with naringin in a mouse model of PD. For neuroprotective effects, 6-hydroxydopamine (6-OHDA) was unilaterally injected into the striatum of mouse brains for a neurotoxin model of PD in the presence or absence of naringin by daily intraperitoneal injection. Our results showed that naringin protected the nigrostriatal DA projection from 6-OHDA-induced neurotoxicity. Moreover, similar to the effects in rat brains, this treatment induced the activation of mammalian target of rapamycin complex 1 (mTORC1), which is well known as an important survival factor for DA neurons, and inhibited microglial activation in the substantia nigra (SN) of mouse brains treated with 6-OHDA. However, there was no significant change of DA phenotypes in the SN and striatum post-treated with naringin compared with 6-OHDA-lesioned mice, despite the treatment being continued for 12 weeks. These results suggest that post-treatment with naringin alone may not be enough to restore the nigrostriatal DA projection in a mouse model of PD. However, our results apparently suggest that naringin is a beneficial natural product to prevent DA degeneration, which is involved in PD.

Carnosic acid protects against 6-hydroxydopamine-induced neurotoxicity in in vivo and in vitro model of Parkinson's disease: involvement of antioxidative enzymes induction.

The neuroprotective effects of carnosic acid (CA), a phenolic diterpene isolated from rosemary (Rosmarinus officinalis), have been widely investigated in recent years, however, its protection in in vivo still unclear. In this study, we investigated the behavioral activity and neuroprotective effects of CA in a rat model of Parkinson's disease (PD) induced by 6-hydroxydopamine (6-OHDA). Rats were treated with 20mg/kg body weight of CA for 3 weeks before 6-OHDA exposure. Results indicated that CA improved the locomotor activity and reduced the apomorphine-caused rotation in 6-OHDA-stimulated rats. Significant protection against lipid peroxidation and GSH reduction was observed in the 6-OHDA rats pretreated with CA. Pretreatment with CA increased the protein expression of γ-glutamate-cysteine ligase catalytic subunit, γ-glutamate-cysteine ligase modifier subunit, superoxide dismutase, and glutathione reductase compared with 6-OHDA-stimulated rats and SH-SY5Y cells. Immunoblots showed that the reduction of the Bcl-2/Bax ratio, the induction of caspase 3 cleavage, and the induction of poly(ADP-ribose) polymerase (PARP) cleavage by 6-OHDA was reversed in the presence of SB203580 (a p38 inhibitor) or SP600125 (a JNK inhibitor) in SH-SY5Y cells. Rats treated with CA reversed the 6-OHDA-mediated the activation of c-Jun NH2-terminal kinase and p38, the down-regulation of the Bcl-2/Bax ratio, the up-regulation of cleaved caspase 3/caspase 3 and cleaved PARP/PARP ratio, and the down-regulation of tyrosine hydroxylase protein. However, BAM7, an activator of Bax, attenuated the effect of CA on apoptosis in SH-SY5Y cells. These results suggest that CA protected against 6-OHDA-induced neurotoxicity is attributable to its anti-apoptotic and anti-oxidative action. The present findings may help to clarify the possible mechanisms of rosemary in the neuroprotection of PD.

Reduction of motor disorder in 6-OHDA-induced severe parkinsonism rats by post treatment with granulocyte-colony stimulating factor.

Granulocyte-colony stimulating factor (G-CSF) induced regeneration of dopaminergic neurons and improved behavior deficit in moderate Parkinson's disease (PD) model mice. Post treatment of G-CSF in severe PD model has not been addressed. A very severe PD model in rats was induced by a high dose 6-hydroxydopamine (6-OHDA) injected into the right medial forebrain bundle to evaluate therapeutic effects of G-CSF. G-CSF (50 microg/kg/day for five days) was given on the 9th day after the 6-OHDA injection. Rotational behavior was examined on the 9th and 28th days. Rats were killed on the 28th day and survival dopaminergic neurons in the substantia nigra, dopaminergic axons and dopaminergic receptor 2 in the striatum were examined. We, for the first time, demonstrated that post treatment with G-CSF reduced abnormal rotational behavior and increased the lesion to non-lesion ratio of dopaminergic fibers in the striatum, but the treatment promoted neither the increase in survival dopaminergic neurons nor the increase in dopaminergic receptor 2 expression. We conclude that post treatment with G-CSF can reduce the abnormal rotational behavior of severe PD rats primarily through relative increases in dopaminergic fibers of the lesion side in the striatum. Results of our study suggest therapeutic potentials of G-CSF for treating severe PD patients.

Effects of low doses of intracerebroventricular 6-OHDA on the levels of monoaminergic neurotransmitters in rat brain structures.

The objective of this study was to determine the degree and specificity of noradrenergic lesions in different areas of the rat brain after intracerebroventricular administration of low doses of 6-hydroxydopamine (6-OHDA) into both lateral ventricles. Our interest focused on the induction of an effective hypothalamic lesion. The results suggest that small doses of 6-OHDA (25 or 50 µg per ventricle) could effectively damage the noradrenergic system in the hypothalamus without significant interfering with the dopamine level and with only a modest reduction in the serotonin concentration.

Effect of prenatal manganese intoxication on [(3)H]glucose uptake in the brain of rats lesioned as neonates with 6-hydroxydopamine.

In the present study we examined the effects of prenatal manganese (Mn) intoxication on [(3)H]glucose uptake in the brain of rats lesioned as neonates with 6-hydroxydopamine (6-OHDA). MnCl(2) . 4H(2)O (10,000 ppm) was added to the drinking water of pregnant Wistar rats for the duration of pregnancy. On the day of parturition, Mn was discontinued as an additive to the drinking water. The control group consisted of rats that consumed water without Mn. Three days after birth, rats in both groups (control and Mn) were pretreated with desipramine hydrochloride (20 mg/kg) and pargyline hydrochloride (50 mg/kg) and injected bilaterally icv with one of three doses of 6-OHDA hydrobromide (15 mug, 30 mug or 67 mug base form in saline on each side) or with saline (control). 6-[(3)H]-D-glucose (500 muCi/kg, ip) was administered to male offspring in adulthood; after 15 min, brain specimens were taken (frontal cortex, hippocampus, striatum, thalamus with hypothalamus, pons and cerebellum) for determination of radioactivity in a liquid scintillation counter. Low dose 6-OHDA (15 mug icv) increased [(3)H]glucose uptake in all brain regions (p < 0.05) in both control and Mn-intoxicated animals. In rats lesioned with a moderate dose of 6-OHDA (30 mug icv), [(3)H]glucose uptake was unaltered in both control and Mn-exposed rats. High dose 6-OHDA (67 mug icv) reduced [(3)H]glucose uptake in all brain regions of Mn-exposed rats (except for cerebellum) compared with the saline group (all, p < 0.05). There was no change in regional brain uptake of [(3)H]glucose in control rats. In conclusion, this study shows that mild neuronal insult (15 mug icv 6-OHDA) increased glucose uptake in the brain while severe damage (concomitant 60 mug icv 6-OHDA and Mn treatment) significantly diminished this process.

[Study on the mechanism of electroacupuncture scalp point penetration therapy in action on apoptosis in the Parkinson's disease rat model].

OBJECTIVE: To explore the mechanism of electroacupuncture scalp point penetration therapy in treatment of the Parkinson's disease (PD). METHODS: Forty Wistar rats were randomly divided into a normal group, a sham-operation group, a model group and an electroacupuncture (EA) group. 6-OHDA was injected into the left striatum to make lateralization PD rat model. Acupuncture at "Baihui" (GV 20)-through-"Taiyang" (EX-HN 5), once each day, 6 days constituting one course. Immunohistochemical method was used to observe the facio-density and the integral optical density of brain-derived neurotrophic factor (BDNF) in the left substantia nigra, and TUNEL method was used to observe the apoptotic amount, and high performance liquid chromatography was used to observe DA contents of the left striatum in each group. RESULTS: As compared with the model group, in the acupuncture group, the facio-density and the integral optical density in the left substantia nigra increased significantly (P < 0.05), the amount of apoptosis decreased significantly (P < 0.05), and the content of striatum DA increased significantly (P < 0.05). CONCLUSION: EA scalp point-through-point therapy may enhance BDNF protein expression level in the substantia nigra to decrease the amount of apoptosis in the PD model rat.

Central or peripheral norepinephrine depletion enhances MK-801-induced plasma corticosterone level in mice.

To investigate the involvement of central or peripheral catecholaminergic systems in the MK-801-induced increase in plasma corticosterone and interleukin-6 levels, we pretreated mice either intracerebroventricularly (i.c.v.) or intraperitoneally (i.p.) with 6-hydroxydopamine (6-OHDA) which depletes catecholamines. Pretreatment of animals with 6-OHDA (50 microg i.c.v. or 100 mg/kg i.p.) significantly enhanced the MK-801 (1 microg i.c.v.)-induced increase in plasma corticosterone level. On the other hand, pretreatment of mice with 6-OHDA (50 microg i.c.v. or 100 mg/kg i.p.) did not affect the MK-801 (1 microg i.c.v.)-induced increase in plasma IL-6 level. These results suggest that central and peripheral catecholaminergic systems are involved in the suppressive regulation of MK-801-induced plasma corticosterone level.

Effect of antidepressant drugs on 6-OHDA-treated mice in the FST.

There is growing evidence suggesting that dopamine could be indirectly involved in the appearance of behavioural effects of antidepressants. In this study, we induced a partial (over 70%) and non-reversible depletion of dopamine-containing neurons in mice by i.c.v. infusion of 6-OHDA. Then, we compared the antidepressant-like effect of drugs (citalopram, paroxetine, desipramine and imipramine) with or without dopamine depletion in the mice forced swimming test. Our results clearly show that lesion with 6-OHDA does not modify the response of mice to desipramine and imipramine, whereas dopamine depletion abolished the antidepressant-like effect of citalopram and paroxetine. It could then be suggested that antidepressant-like effect of selective serotonin reuptake inhibitors (paroxetine and citalopram) in the mice FST requires the activation of dopaminergic pathways to occur.

Trichopus zeylanicus combats fatigue without amphetamine-mimetic activity.

Chronic fatigue is a complex and little understood symptom for which there is no safe and effective pharmacotherapy. The present study was conducted to investigate the effectiveness of Trichopus zeylanicus whole plant powder on fatigue in young Sprague Dawley rats, and aged normal and long-living mutant Ames dwarf mice. Fatigue was evaluated by subjecting the animals to a forced swim test. Trichopus zeylanicus (250 and 500 mg/kg) treated young Sprague-Dawley rats resisted fatigue at a significant level (p < 0.005) compared with controls by an extended swim time in the forced swim test. Oral Trichopus zeylanicus (500 mg/kg) treatment for 2 weeks significantly increased the mobility time in the aged mutant (p < 0.05) and normal mice (p < 0.01) and significantly increased the swim time in the forced swim test in the aged normal mice (p < 0.05). Amphetamine-mimetic activity in Trichopus zeylanicus was excluded by suitable tests. These results show that Trichopus zeylanicus whole plant powder has anti-fatigue effects in young Sprague-Dawley rats and aged normal and mutant Ames dwarf mice providing scientific evidence for the Kani tribal practice in India.

Intravenous 6-hydroxydopamine attenuates vasopressin and oxytocin secretion stimulated by hemorrhage and hypotension but not hyperosmolality in rats.

The present study sought to determine whether chemical destruction of peripheral catecholaminergic fibers with 6-hydroxydopamine (6OHDA) attenuates vasopressin (VP) and oxytocin (OT) secretion stimulated by hemorrhage, hypotension, and hyperosmolality. Rats received 6OHDA (100 mg/kg iv) or vehicle (1 ml/kg iv) on days 1 and 7, and experiments were performed on day 8. Serial hemorrhage (4 samples of 2 ml per 300 g body wt at 10-min intervals) increased plasma VP and OT levels in both groups; however, the increase in plasma VP and OT levels was significantly attenuated in 6OHDA-treated vs. control rats despite a significantly lower mean arterial blood pressure. Similarly, the increase in plasma VP and OT levels in response to hypotension produced by the selective arteriolar vasodilator diazoxide was significantly attenuated in 6OHDA-treated rats. In marked contrast to hemorrhage and hypotension, hyperosmolality produced by an infusion of 1 M NaCl (2 ml/h iv) stimulated increases in plasma VP and OT levels that were not different between 6OHDA-treated and control rats. In a parallel set of experiments, intravenous 6OHDA treatment reduced dopamine--hydroxylase immunoreactivity in the posterior pituitary but had no substantial effect in the hypothalamic paraventricular and supraoptic nuclei. In each experiment, the pressor response to tyramine (250 microg/kg iv) was significantly attenuated in 6OHDA-treated rats, thereby confirming that 6OHDA treatment destroyed sympathetic catecholaminergic fibers. Collectively, these findings suggest that catecholaminergic fibers located outside the blood-brain barrier contribute to VP and OT secretion during hemorrhage and arterial hypotension.

Exogenous and endogenous ghrelin in gastroprotection against stress-induced gastric damage.

Ghrelin, identified in the gastric mucosa has been involved in control of food intake and growth hormone (GH) release but little is known about its influence on gastric secretion and mucosal integrity. The effects of ghrelin on gastric secretion, plasma gastrin and gastric lesions induced in rats by 75% ethanol or 3.5 h of water immersion and restraint stress (WRS) were determined. Exogenous ghrelin (5, 10, 20, 40 and 80 microg/kg i.p.) increased gastric acid secretion and attenuated gastric lesions induced by ethanol and WRS and this was accompanied by the significant rise in plasma ghrelin level, gastric mucosal blood flow (GBF) and luminal NO concentrations. Ghrelin-induced protection was abolished by vagotomy and attenuated by suppression of COX, deactivation of afferent nerves with neurotoxic dose of capsaicin or CGRP(8-37) and by inhibition of NOS with L-NNA but not influenced by medullectomy and administration of 6-hydroxydopamine. We conclude that ghrelin exerts a potent protective action on the stomach of rats exposed to ethanol and WRS, and these effects depend upon vagal activity, sensory nerves and hyperemia mediated by NOS-NO and COX-PG systems.

Long-term stimulation of the subthalamic nucleus in hemiparkinsonian rats: neuroprotection of dopaminergic neurons.

OBJECT: The goal of this study was to evaluate the neuroprotective effects conferred by long-term electrical stimulation of the subthalamic nucleus (STN) against degeneration of dopaminergic neurons by assessing motor functional and immunohistological findings in hemiparkinsonian rats. METHODS: In 13 of 25 rats, a concentric microelectrode was stereotactically implanted into the right STN under the guidance of extracellular microelectrode recording. After this had been done the animals were given an injection of 6-hydroxydopamine (6-OHDA) into the right striatum. Seven of the rats received continuous stimulation (frequency 130 Hz, intensity 80-100 microA) for 2 weeks (Group A); the other six did not receive any stimulation during this period (Group B). Twelve rats did not receive electrode implantation and underwent 6-OHDA injection only; these animals served as a control group (Group C). After 2 weeks, motor function in the rats was evaluated by conducting an amphetamine-induced rotation test. Finally, tyrosine hydroxylase-immunoreactive neurons in the pars compacta of the substantia nigra (SNc) were counted to evaluate the extent of degeneration of dopaminergic neurons. Ipsilateral rotation was significantly decreased in Group A, regardless of the effects of stimulation delivered during the test (p < 0.05). Rats in Group B demonstrated typical circling as did those in Group C, except that on stimulation Group B rats immediately stopped circling or changed direction. Tyrosine hydroxylase-immunoreactive neurons in the SNc were significantly preserved in the animals in Group A, whereas neurons in animals in Groups B and C were moderately depleted (p < 0.01). CONCLUSIONS: Acutely, STN stimulation improved rotation symmetry in rats with moderate SNc degeneration. When STN stimulation had been applied for the preceding 2 weeks, motor function was better and SNc neural degeneration was significantly milder. Subthalamic nucleus stimulation thus appears to protect dopaminergic neurons in this hemiparkinsonian model, in addition to improving motor function in these animals.

The sympathomimetic agent, 6-hydroxydopamine, accelerates cutaneous wound healing.

Using pharmacological stimulation of sympathetic terminals, the present study examines the role of sympathetic post-ganglionic neurons in cutaneous wound healing. Effects of local stimulation of sympathetic post-ganglionic neurons with 0.2 mg/kg 6-hydroxydopamine were studied on the healing of full-thickness skin incisions in rats. Epidermal wound healing was measured by a novel non-invasive quantitative method based on the increasing electrical resistance of healing skin. Dermal healing was determined by measuring wound breaking strength using an Instron Universal Testing device. We report a 35% increase in the rate of epidermal wound healing (P < 0.05. n(e) = 21, n(c) = 18) and a 43% increase in dermal strength (P < 0.05, n(e) = 13, n(c) = 10) after 6-hydroxydopamine treatment. Thus, our results show that pharmacological stimulation of sympathetic post-ganglionic neurons markedly accelerates skin wound healing at both the epidermal and dermal levels. This is the first study to show that peripheral nerve stimulation and specifically sympathetic stimulation accelerates cutaneous wound healing. We discuss these results in relation to neurogenic inflammation.

Dopamine denervation of specific striatal subregions differentially affects preparation and execution of a delayed response task in the rat.

In the present study, the effects of unilateral or bilateral dopamine denervation of either the dorsal or ventral striatum on the preparation and execution of a delayed response task in the rat were investigated. Animals were instructed to hold a lever pressed down by the presentation of a visual and/or acoustic signal, and were required to hold the lever until a trigger stimulus occurred after an unpredictable delay ranging from 2 to 4 s. The trigger stimulus required animals to release the lever and to press a second lever for food reinforcement. The time between instruction and trigger signal represented the preparation phase preceding movement. The motor performance was evaluated by using reaction and movement times in addition to correct responses in each session. Dopaminergic denervation of either the dorsal or ventral striatum ipsilaterally to the side in which the second lever to be pressed was located did not significantly change reaction and movement times, although it reduced the percentage of correct trials. A significant increase of both reaction and movement times was recorded only after bilateral denervation of the ventral striatum. The analysis of incorrect responses indicated that dopaminergic innervation of the two striatal subregions had different functions in the correct execution of the behavioral paradigm. In the group of animals with dorsal lesions the most frequent incorrect response was represented by a lack of the conditioned response to the presentation of the instruction stimulus starting the trial. If the animals reacted properly to this signal, the performance thereafter was correct in the majority of trials. Conversely, animals with ventral lesions exhibited a large repertoire of incorrect responses throughout the paradigm, including premature release or delayed press of levers, and omission of the second lever press. Histological verification of brain coronal sections by tyrosine-hydroxylase immunoreactivity showed that the lesions were confined in either the dorsal or ventral striatum, sparing the lateral region. The data support the hypothesis that dopaminergic innervation enables the two striatal regions to differently participate in the preparation and execution of complex delayed sensorimotor tasks. Indeed, the dorsal striatum seems to be involved in the correct utilization of external sensory information for the initiation of conditioned behavior, whereas, the ventral striatum appears to be mainly concerned with the temporal expectation of impending stimuli that trigger reward-reinforced movements.

Drinking patterns in genetic low-alcohol-drinking (LAD) rats after systemic cyanamide and cerebral injections of THP or 6-OHDA.

A key question related to the role of acetaldehyde and aldehyde adducts in alcoholism concerns their relationship to the genetic mechanisms underlying drinking. Experimentally, the low-alcohol-drinking (LAD) rat represents a standard rodent model having a strong aversion to alcohol. In these experiments, preferences for water vs. alcohol, offered in concentrations from 3% to 30%, were determined over 10 days in adult LAD rats (N = 6 per group). Then a saline vehicle or either 10 or 20 mg/kg of the aldehyde dehydrogenase (AIDH) inhibitor, cyanamide, was injected s.c. twice daily for 3 days. Secondly, either 0.5 or 1.0 microg of tetrahydropapaveroline (THP) was infused i.c.v. twice daily for 3 days in LAD rats (N = 8) and, as a genetic control, THP also was infused identically in Sprague-Dawley (SD) rats (N = 8). The results showed that the lower and higher doses of cyanamide augmented alcohol intakes in 33% and 50% of the LAD rats, respectively, with the patterns of drinking resembling that of genetic high-alcohol-drinking HAD or P rats. Although i.c.v. infusions of THP had little effect on alcohol preference of LAD rats, alcohol drinking was enhanced significantly in the SD rats. In a supplementary study, 200 microg of 6-hydroxydopamine (6-OHDA) also was infused i.c.v. in LAD rats (N = 7) on two consecutive days; no change occurred in the characteristic aversion to alcohol. These findings suggest that in certain individuals, a perturbation in the synthesis of AIDH can modify the genetically based aversion to alcohol, thus precipitating the liability for alcoholism. In that neither THP nor 6-OHDA lesioning exerted any effect on the genetic nondrinking LAD animal suggests that an unknown endogenous factor in the brain must underlie the cyanamide-induced shift to alcohol preference. We conclude that the genetic elements that normally prevent the progression to addictive drinking in most individuals appear to be invariant and irreversible.

Changes in anti-phosphoserine and anti-phosphothreonine antibody binding during the sleep-waking cycle and after lesions of the locus coeruleus.

Cellular responses to many extracellular signals occur through phosphorylation or dephosphorylation of intracellular proteins. To determine whether changes in protein phosphorylation accompany the electrophysiological changes occurring during the sleep-waking cycle, immunocytochemical mapping of cells labeled with anti-phosphoserine and anti-phosphothreonine antibodies was performed on brain sections of sleeping and waking rats. Animals implanted for chronic polysomnographic recordings were sacrificed after either 3h of sleep or 3h of sleep deprivation by gentle handling. Anti-phosphoserine and anti-phosphothreonine staining was mainly localized in neurons and was high in some brain regions, such as cerebral cortex and hypothalamus, and low in others, such as the thalamus. In all cases, the number of cells labeled with either antibody in the cerebral cortex was markedly higher in rats sacrificed after 3h of waking than in rats sacrificed after 3h of sleep. Unilateral lesions of the locus coeruleus by local injection of 6-hydroxydopamine were performed in other animals to determine whether the increase in protein phosphorylation during waking was influenced by the activity of the noradrenergic system, which is higher in waking than in sleep. In animals sacrificed after 3h of spontaneous or forced waking, the number of labeled neurons in the cerebral cortex was decreased on the side in which noradrenergic fibers had been lesioned. These results suggest that 1) neurons exist physiologically in different states of phosphorylation, ranging from a state of very high phosphorylation (e.g., in the cerebral cortex) to a state of very low phosphorylation (e.g., in many thalamic nuclei); 2) the fraction of highly phosphorylated neurons in cerebral cortex is higher in waking than in sleep and 3) part of the immunoreactive phosphorylation present in highly labeled cortical neurons is controlled by the locus coeruleus.

GBR12909 stimulates hypothalamo-pituitary-adrenal activity by inhibition of uptake at hypothalamic dopamine neurons.

We have previously demonstrated that the inhibition of neurotransmitter uptake at dopamine (DA) terminals stimulates the hypothalamo-pituitary-adrenal axis. In the present study we investigated the role of central DA neuronal systems in the regulation of this axis. Administration of the DA uptake inhibitor GBR12909 (3-30 micrograms) into the third ventricle dose-dependently elevated serum adrenocorticotropin hormone (ACTH) levels in rats. GBR12909 (10 micrograms) elevated serum ACTH levels following administration into the paraventricular nucleus of the hypothalamus but not into the lateral ventricle. The administration of 6-OHDA into the third ventricle significantly decreased DA content in the hypothalamus and striatum and significantly attenuated the ACTH response to GBR12909 (10 micrograms, third ventricle or 10 mg/kg, i.p.). Conversely, 6-OHDA lesions of the medial forebrain bundle, which depleted 99% of DA in the caudate but did not decrease DA content in the hypothalamus, and did not attenuate the ACTH response to i.p. GBR12909. Measurement of GBR12909-induced inhibition of [3H]DA uptake into synaptosomal preparations indicates the presence of GBR12909-sensitive DA transporters in the region of the hypothalamus surrounding the third ventricle. The present findings suggest that an endogenous DA neuronal system terminating in the hypothalamus mediates the effects of stimulants on hypothalamo-pituitary-adrenal function and might play a role in the ongoing regulation of hypothalamo-pituitary-adrenal activity.