(6R)-L-erythro-5,6,7,8-Tetrahydrobiopterin (BH4) is an essential cofactor for tyrosine hydroxylase (TH), tryptophan hydroxylase, phenylalanine hydroxylase, and nitric-oxide synthase. These enzymes synthesize neurotransmitters, e.g. catecholamines, serotonin, and nitric oxide (NO). We established mice unable to synthesize BH4 by disruption of the 6-pyruvoyltetrahydropterin synthase gene, the encoded protein of which catalyzes the second step of BH4 biosynthesis. Homozygous mice were born at the almost expected Mendelian ratio, but died within 48 h after birth. In the brain of homozygous mutant neonates, levels of biopterin, catecholamines, and serotonin were extremely low. The number of TH molecules was highly dependent on the intracellular concentration of BH4 at nerve terminals. Alteration of the TH protein level by modulation of the BH4 content is a novel regulatory mechanism. Our data showing that catecholaminergic, serotonergic, and NO systems were differently affected by BH4 starvation suggest the possible involvement of BH4 synthesis in the etiology of monoamine-based neurological and neuropsychiatric disorders.Catecholamines, e.g. dopamine (DA), 1 norepinephrine (NE), and epinephrine, and serotonin (5-hydroxytryptamine; 5HT) are well known to be involved in many aspects of brain function, including mood, addiction, reward, and sleep. Alteration in the metabolism of these monoamines leads to abnormalities in behavior and neuropsychiatric states. For example, increased dopaminergic neurotransmission in the mesolimbic system induces hallucinations, illusions, and delusions, which are the major symptoms of schizophrenia (1). Depletion of monoamines may be the cause of depression, because drugs that can increase the level of monoamines and blockers of norepinephrine or serotonin uptake show therapeutic effects in patients with depression (2). Abnormal monoamine metabolism is also suspected to be involved in Tourette's syndrome, obsessive-compulsive disorder, Rett syndrome, and infantile autism (3, 4).Biosynthesis of catecholamines and serotonin is mainly regulated by the activity of the hydroxylation reaction catalyzed by tyrosine hydroxylase (TH) in catecholamine synthesis (5) and by tryptophan hydroxylase (TPH) in 5HT synthesis (6). The TH activity is tightly regulated by several factors (for review, see Ref. 7 and references therein). Short term regulation is achieved by phosphorylation of the enzyme and by feedback inhibition with the end products. Long term regulation is mainly governed at the transcriptional level.(6R)-L-erythro-5,6,7,8-Tetrahydrobiopterin (BH4) is an essential cofactor for both TH and TPH. Although many cofactors such as folic acid and cobalamin (vitamin B12) are required to be dietary supplemented, mammals have a de novo biosynthetic pathway for BH4. In this pathway, guanosine triphosphate (GTP) is converted to 7,8-dihydroneopterin triphosphate by GTP cyclohydrolase 1 (GCH), and 7,8-dihydroneopterin triphosphate is then converted to 6-pyruvoyltetrahydropterin by pyruvoyltetrahydropteri...

Aromatic-L-amino-acid decarboxylase (AADC) is an enzyme that plays an essential role in synthesizing catecholamines and serotonin in neuronal and endocrine tissues. AADC has also been detected in other nonneuronal tissues including liver and kidney, although its physiological role in nonneuronal tissues has not yet been defined. Previously we have cloned a human AADC cDNA from a neuronal tissue (pheochromocytoma) [Ichinose, H., Kurosawa, Y., Titani, K., Fujita, K., & Nagatsu, T. (1989) Biochem. Biophys. Res. Commun. 164, 1024-1030] and the corresponding genomic DNA [Sumi-Ichinose, C., Ichinose, H., Takahashi, E., Hori, T., & Nagatsu, T. (1992) Biochemistry 31, 2229-2238]. Here we present isolation and characterization of AADC cDNA and genomic DNA from a nonneuronal tissue (human liver). The nonneuronal and neuronal AADC mRNAs differed only in the region corresponding to the untranslated first exon. The first exon for the nonneuronal-type mRNA was located 4.2 kilobases upstream to that for the neuronal-type mRNA and 22 kilobases from exon 2, to which it is spliced. Determination of the transcription initiation site indicated that the length of the nonneuronal-type exon 1 was 200 bp. A TATA box-like motif was located between positions -26 and -20 from the transcription initiation site. These results showed that an alternative usage of the first exon in the 5'-untranslated regions produces two types of mRNAs in AADC and suggested that alternative splicing would regulate the tissue-specific expression of AADC.

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The physiological relationship between thyroid hormone and the catecholamines has been investigated for many years (1, 2). In spite of numerous studies it is still not clear whether thyroid status alters the sensitivity of the heart to catecholamines. Coville (3) has reported that thyroid hormone increased the magnitude of the response to catecholamines, while some authors have failed to show that altering the thyroid status affects the response to catecholarnines (4, 5). Recently, it was tried to explore some effects of thyroidectomy on the response of alpha and beta adrenergic receptor mechanisms to catecholamines (6). A few workers have reported the existence of both alpha and beta receptors in the hearts of mammals (7,8).
Present studies reported here were made in an attempt to clarify some of the discrepancies in many reports by seeking a quantitative difference between different types of cardiac adrenergic receptor mechanisms after 6-propyl-2-thiouracil (PTU) treatment. METHODSMale Wistar strain rats (170 to 200 g) were used for experiments. The rats were divided into two groups. Group A was served as control. Group B -,vas in a hypothyroid state produced by feeding on 0.15 % PTU diet for 6 to 8 weeks. This group developed dryness of fur, slow heart rate and a great retardation in growth.Each rat was killed by a blow on the head and the heart was rapidly removed and the left atrium dissected free. Isolated left atria were suspended in a 25-m1 tissue bath containing Ringer solution of the following composition (mM); NaCl, 158; KCI, 5.6; CaCl2, 3.6; Na HCO2, 3.6; and glucose, 15.6. The solution was maintained at a temperature of 30 -:---0.5 C and saturated with pure oxygen. The preparations were driven at a rate of 180 beats/min.through an electrode by a square wave pulse of 6 msec duration and voltage of approximately twice threshold delivered by a stimulator (Nihon Kohden MSE-3). A force displacement transducer (Nihon Kohden SB-IT) was used for measurement of isometric contractile force which was recorded by an ink writing oscillograph. A resting tension of 0.5 g was placed on each atrium. The preparations were allowed to equilibrate in Ringer solution for about 60 min. before exposing them to drugs. Drugs were dissolved before use in 0.9;V saline.The drugs used in these experiments were 1-phenylephrine hydrochloride , 1-isoproterenol