Robert A. Mueller

Adrenal Tyrosine Hydroxylase: Compensatory Increase in Activity after Chemical Sympathectomy

Destruction of peripheral sympathetic nerve endings with 6-hydroxydopamine causes a disappearance of cardiac tyrosine hydroxylase, accompanied by a twofold increase in adrenal tyrosine hydroxylase and a small increase in phenyl-ethanolanine-N-methyl transferase. No change in adrenal catecholamine content occurs under these conditions.

Increased Tyrosine Hydroxylase Activity after Drug-induced Alteration of Sympathetic Transmission

AFTER the destruction of sympathetic nerve endings by 6-hydroxydopamine1,2, there is a compensatory increase in the catecholamine synthesis and tyrosine hydroxylase activity in the adrenal gland3. We have examined the actions of reserpine and phenoxybenzamine on adrenal tyrosine hydroxylase activity. These drugs, like 6-hydroxydopamine2, interfere with postganglionic sympathetic transmission, but by different mechanisms4–6. We also studied the alterations of this enzyme in an embryologically related tissue, the superior cervical sympathetic ganglion. Reserpine and phenoxybenzamine increased the activity of adrenal tyrosine hydroxylase, and reserpine also increased the activity of tyrosine hydroxylase in the superior cervical ganglion. This increase in enzyme activity was prevented by interruption of nerve impulses by decentralization.

EVIDENCE FOR INVOLVEMENT OF 5-HYDROXYTRYPTAMINE IN THE ACTIONS OF AMPHETAMINE

1 Pargyline treatment, 1 h before (+)‐amphetamine (1 mg/kg), reduced amphetamine‐stimulated motor activity. This inhibition was reversed in animals pretreated with p‐chlorophenylalanine (PCPA). 2 Following treatment with PCPA or 5,6‐dihydroxytryptamine (5,6‐DHT), amphetamine‐induced locomotor activity was significantly potentiated. The increased response to amphetamine in PCPA‐treated rats was reversed in animals pretreated with 5‐hydroxytryptophan. 3 The inhibition of amphetamine‐stimulated locomotor activity by treatment with 6‐hydroxydopamine was not reversed by PCPA treatment. 4 Stereotypies produced by amphetamine were not found to be altered by depletion of 5‐hydroxytryptamine. 5 Induction of adrenal tyrosine hydroxylase activity produced by chronic amphetamine administration was significantly potentiated by PCPA, emphasizing the involvement of a 5‐hydroxytryptamine inhibitory system in more than one action of amphetamine.

Neonatal and adult 6-hydroxydopamine-induced lesions differentially alter tachykinin and enkephalin gene expression

Abstract: The present investigation examined the effects of neonatal and adult 6‐hydroxydopamine (6‐OHDA)‐induced lesions of dopaminergic neurons on opioid and tachykinin peptides and their gene expression in the rat basal ganglia. This work was undertaken to determine if changes in these neuropeptide systems were contributing to the differing behavioral responses observed between neonatally and adult‐lesioned rats after dopamine agonist administration. [Met5]Enkephalin (ME) content was increased in striatal tissue from both 6‐OHDA‐lesioned groups when compared with unlesioned controls. Dynorphin‐A (1–8) content was not altered by the 6‐OHDA lesions. The tachykinin peptides substance P and neurokinin A were significantly decreased in level in the striatum and substantia nigra of neonatally lesioned rats, but not in the adult‐lesioned rats, when compared with unlesioned controls. Proenkephalin mRNA abundance (quantified by an RNA‐cDNA hybridization technique) and precursor level (as reflected by cryptic ME content) were increased in the striatum of both neonatally and adult‐lesioned rats. The abundance of preprotachykinin mRNA coding for the tachykinin peptides was markedly decreased in the neonatally lesioned rats, whereas only a small reduction was observed in the adult‐lesioned rats. These results suggest that destruction of dopamine‐containing terminals with 6‐OHDA elevates the level of ME by accelerating transcriptional and/or translational processes; conversely, the reduced content of tachykinins in neonatally lesioned rats may be due to a reduction in such processes. Thus, preproenkephalin‐A and preprotachykinin gene expression are differentially regulated after lesioning of catecholamine‐containing neurons, an observation suggesting a close functional relationship among these neurotransmitter systems. Furthermore, of the peptides studied, only levels of the tachykinin peptides were differentially altered in the striatum and substantia nigra of the neonatally lesioned rats compared with adult‐lesioned rats; therefore, these peptides may be associated with the distinctive behavioral differences between neonatally and adult 6‐OHDA‐lesioned rats given dopamine agonists.

Clinical Studies of Methylphenidate Serum Levels in Children and Adults

Abstract Although methylphenidate (MPH) is used widely in the treatment of hyperactive children, and although it seems to be a safe and effective drug, very little is known about the pharmacology, pharmacokinetics, and metabolism of the drug, and the relationship between serum levels and subtle side effects. We report on the progress in applying a reliable and sensitive gas chromatography-mass spectrometric assay for MPH in serum to clinical studies of hyperactive children and adults. Although drug half-life in all subjects is remarkably constant (2 to 4 hours), there is a wide range of inter-individual variation in serum levels and, more disturbing, a wide range of intra-individual variations in serum levels. The clinical importance of this work is discussed.

Metabolic mapping of the rat brain after subanesthetic doses of ketamine : potential relevance to schizophrenia

Subanesthetic doses of ketamine have been shown to exacerbate symptoms in schizophrenia and to induce positive, negative, and cognitive schizophrenic-like symptoms in normal subjects. The present investigation sought to define brain regions affected by subanesthetic doses of ketamine, using high resolution autoradiographic analysis of 14C-2-deoxyglucose (2-DG) uptake and immunocytochemical staining for Fos-like immunoreactivity (Fos-LI). Both functional mapping approaches were used because distinct and complementary information is often obtained with these two mapping methods. Ketamine, at a subanesthetic dose of 35 mg/kg, substantially increased 2-DG uptake in certain limbic cortical regions, including medial prefrontal, ventrolateral orbital, cingulate, and retrosplenial cortices. In the hippocampal formation, the subanesthetic dose of ketamine induced prominent increases in 2-DG uptake in the dentate gyrus, CA-3 stratum radiatum, stratum lacunosum moleculare, and presubiculum. Increased 2-DG uptake in response to 35 mg/kg ketamine was also observed in select thalamic nuclei and basolateral amygdala. Ketamine induced Fos-LI in the same limbic cortical regions that exhibited increased 2-DG uptake in response to the subanesthetic dose of the drug. However, no Fos was induced in some brain regions that showed increased 2-DG uptake, such as the hippocampal formation, anterioventral thalamic nucleus, and basolateral amygdala. Conversely, ketamine induced Fos in the paraventricular nucleus of the hypothalamus and central amygdala, although no effect of the drug on 2-DG uptake was apparent in these regions. In contrast to the increase in 2-DG uptake observed in select brain regions after the subanesthetic dose, an anesthetic dose of ketamine (100 mg/kg) produced a global suppression of 2-DG uptake. By contrast, a robust induction of Fos-LI was observed after the anesthetic dose of ketamine that was neuroanatomically identical to that produced by the subanesthetic dose. Results of the present investigation show that anesthetic and subanesthetic doses of ketamine have pronounced effects on regional brain 2-DG uptake and induction of Fos-LI. The alterations in regional brain metabolism induced by the subanesthetic dose may be relevant to effects of ketamine to induce schizophrenic-like symptoms.

Serotonergic innervation of the rat caudate following a neonatal 6-hydroxydopamine lesion: an anatomical, biochemical and pharmacological study.

6-Hydroxydopamine (6-OHDA) treatment of neonatal rats resulted in a dose-related loss of striatal dopamine (DA). These reductions corresponded closely with the loss of tyrosine hydroxylase-containing terminals at this brain site. Striatal serotonin (5-HT) concentration increased only after DA was maximally depleted by the highest dose of 6-OHDA. Quantitative immunohistochemistry revealed that the increased 5-HT content after neonatal 6-OHDA lesioning was due to a proliferation of 5-HT nerve terminals. The density of immunoreactive 5-HT-containing terminals appeared to increase more than did the 5-HT content. The present study examined whether 5-HT hyperinnervation was playing a role in behavioral responses induced by D1-DA agonists and antagonists in neonatally lesioned rats, because reports have suggested that these drugs may interact with 5-HT receptors. However, SCH-23390, the D1-DA antagonist (0.3 mg/kg), did not alter behavioral responses to 5-HTP and SKF-38393 (3 mg/kg), a D1-DA agonist did not produce any signs of activating 5-HT receptors in 5,7-DHT-lesioned rats. These data indicate that these compounds affecting D1-DA receptors do not have a significant effect on 5-HT function at doses which have maximal effects on D1-DA receptor function. Pretreatment with the 5-HT antagonist methysergide did not produce a change in apomorphine-induced locomotion and did not antagonize the self-mutilation or the other behaviors produced by L-DOPA or SKF-38393 in neonatally lesioned rats, suggesting that 5-HT hyperinnervation is not responsible for these drug-induced changes in neonatal 6-OHDA-lesioned rats.

SCH-23390 antagonism of a D-2 dopamine agonist depends upon catecholaminergic neurons

SCH-23390, a selective D-1 dopamine antagonist, was found to antagonize the locomotor stimulation induced by LY-171555, an action similar to that for haloperidol in control animals. However, this action of SCH-23390 was prevented in rats treated with 6-hydroxydopamine (6-OHDA) or with reserpine plus alpha-methyl-tyrosine pretreatment. These results indicate that the action of SCH-23390 to antagonize D-2 dopamine receptor actions is dependent upon functional catecholamine-containing neurons. In contrast to the lack of action of SCH-23390 to antagonize LY-171555 in 6-OHDA-treated rats, SCH-23390 blocked the locomotor stimulation induced by SKF-39393, a D-1 dopamine agonist, after this treatment. Thus, D-1 dopamine receptors are distinct from D-2 receptor sites and can exhibit a behavior similar to that observed when D-2 receptors are stimulated. These data suggest that D-1 receptor sites modulate D-2 dopamine receptor function through a mechanism dependent upon functionally intact catecholamine-containing neurons.

Postconditioning Prevents Reperfusion Injury by Activating δ-Opioid Receptors

Background:While postconditioning has been proposed to protect the heart by targeting the mitochondrial permeability transition pore (mPTP), the detailed mechanism underlying this action is unknown. The authors hypothesized that postconditioning stimulates opioid receptors, which in turn protect the heart from reperfusion injury by targeting the mPTP. Methods:Rat hearts (both in vivo and in vitro) were subjected to 30 min of ischemia and 2 h of reperfusion. Postconditioning was elicited by six cycles of 10-s reperfusion and 10-s ischemia. To measure nitric oxide concentration, cardiomyocytes loaded with 4-amino-5-methylamino-2′,7′-difluorofluorescein were imaged using confocal microscopy. Mitochondrial membrane potential was determined by loading cardiomyocytes with tetramethylrhodamine ethyl ester. Results:In open chest rats, postconditioning reduced infarct size, an effect that was reversed by both naloxone and naltrindole. The antiinfarct effect of postconditioning was also blocked by the mPTP opener atractyloside. In isolated hearts, postconditioning reduced infarct size. Morphine mimicked postconditioning to reduce infarct size, which was abolished by both naltrindole and atractyloside. N-nitro-l-arginine methyl ester and guanylyl cyclase inhibitor 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one blocked the action of morphine. Further experiments showed that morphine produces nitric oxide in cardiomyocytes by activating &dgr;-opioid receptors. Moreover, morphine could prevent hydrogen peroxide–induced collapse of mitochondrial membrane potential in cardiomyocytes, which was reversed by naltrindole, N-nitro-l-arginine methyl ester, and the protein kinase G inhibitor KT5823. Conclusions:Postconditioning protects the heart by targeting the mPTP through activation of &dgr;-opioid receptors. The nitric oxide–cyclic guanosine monophosphate–protein kinase G pathway may account for the effect of postconditioning on the mPTP opening.

Inhibition of neuronally induced tyrosine hydroxylase by nicotinic receptor blockade

Abstract The increase in tyrosine hydroxylase activity in the rat superior cervical ganglion produced by reserpine is abolished by decentralization or administration of the nicotinic blockers chlorisondamine and pempidine. In contrast, the rise in adrenal tyrosine hydroxylase activity is neither inhibited by nicotinic nor muscarinic receptor blockers, although it is abolished by splanchnic transection. Since chlorisondamine and pempidine diminish the magnitude of adrenal catecholamine depletion produced by reserpine to the same extent as denervation, it is unlikely that incomplete blockade of the nicotinic receptors is responsible for the inability to block the increase in adrenal enzyme activity. It is possible that in the adrenal medulla either another cholinergic receptor or another neurotransmitter is involved in mediating the trans-synaptic increase in tyrosine hydroxylase. Pempidine, chlorisondamine and atropine all produce an increase in adrenal tyrosine hydroxylase when used alone. This can be prevented by prior splanchnic transection.