Mitsuaki Suda

α-Human atrial natriuretic polypeptide is released from the heart and circulates in the body

In order to clarify whether or not atrial natriuretic polypeptides are hormones in man, we have measured plasma alpha-human atrial natriuretic polypeptide (alpha-hANP)-like immunoreactivity (alpha-hANP-LI) with or without extraction procedure. alpha-hANP-LI was detected in plasma extracts from all 5 normal subjects and 7 patients with heart diseases. The alpha-hANP-LI concentration in normal peripheral plasma was 37.7 +/- 7.0 pg/ml (mean +/- SE). Plasma concentrations of alpha-hANP-LI in the coronary sinus obtained by cardiac catheterization were 3 to 10 times higher than those in the peripheral vein, inferior vena cava, right atrium, pulmonary artery and aorta. High performance gel permeation chromatography coupled with a radioimmunoassay (RIA) for alpha-hANP revealed that alpha-hANP-LI in normal peripheral plasma eluted at the position corresponding to that of authentic alpha-hANP without detectable amounts of high molecular weight forms. alpha-hANP-LI extracted from plasma taken from the coronary sinus of two patients also showed a single peak of alpha-hANP-LI co-eluting with alpha-hANP. In contrast, not only alpha-hANP but gamma-hANP and beta-hANP, high molecular weight forms, were present in the human atrial tissue. These results indicate that alpha-hANP is the predominant form of alpha-hANP-LI in human plasma and that this form generated in the atrial cardiocytes is preferentially released from these cells and circulates in the body.

Occurrence of atrial natriuretic polypeptide in brain

In order to determine whether or not atrial natriuretic polypeptides (ANPs) exist in the brain, we have studied extracts from the rat brain using a specific radioimmunoassay (RIA) for alpha-atrial natriuretic polypeptide. The presence and widespread distribution of alpha-rat ANP-like immunoreactivity (alpha-rANP-LI) have been demonstrated in the rat brain. The highest concentration of alpha-rANP-LI (20-22 ng/g) is in the hypothalamus and the septum. Moderate concentrations of alpha-rANP-LI (2-8 ng/g) are also found in the midbrain, cerebral cortex, olfactory bulb, thalamus, pons-medulla and hippocampus. High performance gel permeation chromatography coupled with the RIA revealed that alpha-rANP-LI found in the rat brain consists of three components eluting at the positions of gamma-rat ANP (gamma-rANP), beta-rat ANP (beta-rANP) and alpha-rANP, respectively. Among these a low molecular weight form of alpha-rANP-LI emerging at the elution position corresponding to alpha-rANP is predominant in the rat brain. This is in contrast to the finding that gamma-rANP, a high molecular weight form of 13k daltons is the dominant form of alpha-rANP-LI in the rat atrium. These results clearly show that there exists a widespread neural system containing ANPs in the brain.

Radioimmunoassay for α-human and rat atrial natriuretic polypeptide

Abstract Using a synthetic common carboxy-terminal fragment of α-human atrial natriuretic polypeptide (α-hANP) and α-rat atrial natriuretic polypeptide (α-rANP), we have produced an antiserum for α-ANP(17–28) and established a radioimmunoassay (RIA) for α-ANP that recognizes α-hANP and α-rANP equally. High performance gel permeation chromatography coupled with the RIA revealed that α-hANP-like immunoreactivity (α-hANP-LI) in the human atrium consists of three major components, γ-hANP, α-hANP and another. On the other hand, α-rANP-LI in the rat atrium comprised at least two components, 13K α-rANP-LI and 3K–5K α-rANP-LI, which were presumably γ-rANP and β-rANP, respectively. Thus, considerable amounts of γ-hANP and γ-rANP are present in human right auricles and rat atria, respectively. The RIA established in this study provides a useful tool to investigate the pathophysiological significance of α-ANP and related peptides in cardiovascular disorders and shows that γ-ANP is not only a precursor of α-ANP but acts as an important hormone.

Existence of atrial natriuretic polypeptide in kidney

Using a specific radioimmunoassay (RIA) for alpha-atrial natriuretic polypeptide (alpha-ANP), we have demonstrated the presence of alpha-rat ANP-like immunoreactivity (alpha-rANP-LI) in the rat kidney which is considered to be a target organ for atrial natriuretic polypeptides released from the heart. Most of alpha-rANP-LI was localized in the cortex. High performance gel permeation chromatography coupled with the RIA revealed that renal alpha-rANP-LI was eluted at the position of a low molecular weight form corresponding to alpha-rANP without detectable amounts of high molecular weight forms. This is in contrast to the observation that gamma-rANP, a high molecular weight form of 13k daltons, is the dominant form of alpha-rANP-LI in the rat atrium. In water-deprived rats, the concentration and content of alpha-rANP-LI in the kidney showed a significant decrease compared with control rats. In addition, the alpha-rANP-LI concentration and content in this organ revealed a substantial decrease after perfusion with physiological saline. These results indicate the existence of atrial natriuretic polypeptide (ANP) in the kidney and suggest that part of renal ANP may originate from the heart.

Leumorphin is a novel endogenous opioid peptide derived from preproenkephalin B

Using synthetic leumorphin, we obtained antisera for leumorphin and set up two radioimmunoassays (RIAs) with different specificities. Gel exclusion chromatography coupled with the two RIAs showed the existence of a considerable amount of leumorphin-like peptide in water extracts from porcine neuro-intermediate pituitaries. Reverse phase high performance liquid chromatography revealed that leumorphin-like peptide in the water extracts was indistinguishable from synthetic leumorphin. These results along with potent opioid activity of leumorphin indicate that leumorphin is a novel endogenous opioid peptide derived from preproenkephalin B.

A novel opioid peptide, leumorphin, acts as an agonist at the κ opiate receptor

Abstract The primary structure of the common precursor of porcine β-neo-endorphin and dynorphin (preproenkephalin B) has shown the existence of a third leucine-enkephalin (leu-enkephalin) sequence with a C-terminal extension of 24 amino acids. This nonacosapeptide, named leumorphin, was approximately 70 times more potent than leuenkephalin in inhibiting the contraction of the myenteric plexus-longitudinal muscle preparation of the guinea pig ileum. This action of leumorphin, like those of β-neo-endorphin and dynorphin, was antagonized less effectively by naloxone than that of leuenkephalin, but more effectively by Mr2266, an antagonist relatively specific for the κ type opiate receptor. The inhibitory action of leumorphin or β-neo-endorphin on the contraction of the guinea pig ileum muscle strip was reduced in a dose-dependent manner by pretreatment with dynorphin and vice versa . Leumorphin as well as β-neo-endorphin and dynorphin inhibits the contraction of the rabbit vas deferens which is known to have only the κ type opiate receptor. This action was also effectively antagonized by Mr2266. It is concluded that leumorphin has potent opioid activity and acts at the κ receptor, like other opioid peptides derived from preproenkephalin B.

Biosynthesis and distribution of opioid peptides

Group III opioid peptides are derived from proenkephalin B. The processing of this precursor peptide is still only partly understood and we still do not know how many final products come from proenkephalin B and whether Leu-enkephalin is produced from Group III peptides. However, potent biologic activity of Group III opioid peptides and existence of opiate-receptors specific for these peptides (k receptors) strongly suggest that Group III opioid peptides are not precursors of Leu-enkephalin but bioactive substances per se. Further studies should clarify details of the processing of proenkephalin B.

Accelerated Natriuresis Induced by Synthetic Atrial Natriuretic Polypeptide in Spontaneously Hypertensive Rats

Effects of synthetic alpha human atrial natriuretic polypeptide (alpha-hANP) on diuresis, natriuresis and mean arterial blood pressure (MAP) were compared between 4-5 month-old male spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY) under ether anesthesia. In both groups, the peptide injected (0.5 micrograms/100g body weight, i.v.) caused potent (about ten fold), rapid and short-acting (for 15 min) increases in sodium (Na+) and chloride excretions and also an increase in urine flow and potassium excretion with lesser magnitude. Although ratios of the maximum response to basal value were much the same, net increases in urine flow and Na+ output were significantly greater in SHR than in WKY. As to the effect on MAP, a rapid (within 2 min) fall observed in the two groups. These results suggest that the atrial natriuretic peptide may be involved in the altered regulatory mechanism of fluid and electrolyte balance in SHR models with genetic hypertension.

Changes in the immunoreactivities of an opioid peptide leumorphin in the hypothalamus and anterior pituitary during the estrous cycle of the rat and their relation to sexual behavior

Leumorphin, an opioid peptide whose functions are unknown, is found in mammalian brain and pituitary and stimulates lordosis behavior in estrogen-treated female rats. To elucidate the role of leumorphin in the physiological control of female sexual behavior, the levels of immunoreactive (ir) leumorphin as well as ir dynorphin (dynorphin A) were measured in the rat brain and pituitary during the estrous cycle. There was a clear variation of ir leumorphin in the hypothalamus and anterior pituitary during the estrous cycle. The levels of ir leumorphin in the hypothalamus and anterior pituitary on the afternoon of proestrus were significantly higher (P less than 0.01) than those on the afternoons of estrus and metestrus. The rise in the hypothalamic levels of ir leumorphin on the afternoon of proestrus was correlated with the receptivity of lordosis during the estrous cycle. Furthermore, there was a close correlation with ir dynorphin levels. These findings are in agreement with studies demonstrating a common precursor for leumorphin and dynorphin. Ir leumorphin in the hippocampus and neurointermediate pituitary did not change significantly during the estrous cycle. Because the leumorphin antiserum used recognizes rimorphin (dynorphin B) 1.78 times more than porcine leumorphin on a molar basis, high performance-gel permeation chromatography was done on pooled extracts of hypothalamus taken at proestrus and estrus. The peak in the leumorphin-like substance in the activation of sexual behavior is discussed.

Parallel distribution of methionine-enkephalin-arg6-gly7-leu8 with methionine-enkephalin, leucine-enkephalin and methionine-enkephalin-arg6-phe7 in human and bovine brains

Using specific radioimmunoassays(RIAs) for methionine-enkephalin(Met-Enk), leucine-enkephalin(Leu-Enk), methionine-enkephalin-Arg6-Gly7-Leu8 (Met-Enk-Arg-Gly-Leu) and methionine-enkephalin-Arg6-Phe7 (Met-Enk-Arg-Phe), we studied the regional distribution of these opioid peptides in human and bovine brains. Met-Enk-Arg-Gly-Leu was distributed in parallel with Met-Enk, Leu-Enk and Met-Enk-Arg-Phe in human and bovine brains. The ratios of molar concentrations of these peptides are almost constant in various regions of human and bovine brains and similar to the ratio of these peptides contained in preproenkephalin A. Gel exclusion chromatography and HPLC coupled with respective RIAs showed the existence of authentic peptides without any detectable high molecular weight forms. These results indicate the parallel distribution of Met-Enk-Arg-Gly-Leu with Met-Enk, Leu-Enk and Met-Enk-Arg-Phe in various regions of human and bovine brains and further suggest that these opioid peptides are derived from the same precursor as that in the adrenal medulla and that the processing of preproenkephalin A is almost complete in human and bovine brains.