novel cardiac syndrome known as 'tako-tsubo' cardiomyopathy or 'transient left ventricular apical ballooning' and which mimics acute myocardial infarction has recently been proposed by Japanese cardiologists. 1,2 It is characterized by (1) acute onset and reversible left ventricular (LV) apical wall motion abnormalities (ballooning) with chest symptoms, (2) electrocardiographic (ECG) changes (ie, ST elevation), (3) minimal myocardial enzymatic release, and (4) no significant stenosis on coronary angiography. Emotional or physical stress usually precedes the attack, but the precise etiology remains to be elucidated. Myocardial ischemia -reperfusion because of microvascular spasm or a direct cardiac effect of catecholamines might be the cause of this transient abnormality of LV wall motion, yet clinical approaches have been limited in detecting its pathogenesis. To understand the causal relation between stress and this syndrome, investigations using a whole animal model are fundamental. We previously reported that immobilization (IMO) of the rat, a useful animal model of emotional stress, 3 produced reversible elevation of the ST segment on ECG, which was normalized by pretreatment with -adrenoceptor blockers, but not by potent coronary vasodilators such as a calcium channel blocker or nitroglycerin. 4 In the present study, we used left ventriculography (LVG) in the rat to examine stress-induced abnormalities of LV wall motion.A polyethylene catheter (PE50, Becton Dickinson, Sparks, MD, USA) was inserted into the left ventricle via the common carotid artery of adult female Wistar rats (n=10; 250-300 g). Under anesthesia induced with pentobarbital (40 mg/kg), 1 ml of contrast medium (Iopamidol 370, Schering Ag, Berlin, Germany) was injected and LVG was performed (controls) using the digital subtraction system (DSA) (SERIES 9600, OEC Medical Systems Inc, Salt Lake City, UT, USA). LVG was recorded at 30 frames/s, and evaluated independently by 2 observers. The margins of the LV images were traced, and the end-systolic or end-diastolic LV areas were also evaluated using the NIH image. Percent contraction was determined as 100 × (end-diastolic area -end-systolic area) / end-diastolic area. Statistical analysis was performed by one-way ANOVA followed by Fisher's protected least significant difference test, using StatView software (Abacus Concepts, Berkeley, CA, USA).The next day, the rats were exposed to IMO stress for up to 30 min by securing the rat on its back to a board using adhesive tape, and LVG was performed without anesthesia. All animal manipulations were approved by Wakayama Medical University Animal Care and Use Committee.In response to stress, generalized LV hypokinesis was observed in 4 cases and apical ballooning was observed in 4 cases (Fig 1). The other 2 cases were judged to be unaffected by stress (Fig 2). The following day, rats that had shown LV hypokinesis (n=3) or apical ballooning (n=3) were again subjected to IMO stress after pretreatment with an -adrenoceptor blocker, amosulalol hydrochloride (...

INTRODUCTIONRecent clinical studies (1-4) have revealed the clinical features of 'Tako-tsubo (Ampulla)' cardiomyopathy mimicking acute myocardial infarction. This is characterized by: (i) acute onset, reversible left ventricular (LV) apical wall motion abnormalities (ballooning) with chest symptoms; (ii) electrocardiographic (ECG) changes (ST elevation); (iii) minimal myocardial enzymatic release; and (iv) no significant stenosis or spasm on coronary angiography. The backgrounds of these patients are unique, but have the following two characteristics: first, there is an emotional or physical stress event triggering the attack (33-86%); second, the patient is often an elderly (postmenopausal) female (75-93%). The precise etiology of this syndrome is yet to be determined, so finding a similar animal model would be extremely valuable for the study of the underlying pathological mechanisms of this condition. We have produced this syndrome by immobilization stress (IMO) in rats (5); this produces a reversible elevation of the ST segment on the ECG as well as the reversible LV apical ballooning in the left ventriculography (LVG), both of which were normalized by pretreatment with αβ-adrenoceptor blockers (6,7). In order to study whether these cardiac changes are especially predominant in elderly female subjects (8), we investigated LVG and ECG under IMO in ovariectomized (animal model of post-menopause) and estradiol-supplemented ovariectomized female rats. METHODAdult (24-to 32-week-old) female Wistar rats weighing 250-300 g, (n = 16) were purchased from Kiwa Laboratories (Wakayama, Japan). Estradiol 10 mg (β-estradiol; Sigma, St. Louis, MO, U.S.A.) was introJournal of Cardiovascular Pharmacology™ 42 (Suppl. 1):S117-S119 Summary: Reduction of estrogen levels may underlie the high incidence of 'Tako-tsubo (Ampulla) cardiomyopathy in postmenopausal females. Ovariectomized (OVX) and estradiol-supplemented ovariectomized female rats (OVX + E) were subjected to immobilization stress, an animal model of Tako-tsubo cardiomyopathy. In order to evaluate cardiac changes, left ventriculography and electrocardiography were performed under anesthesia (control). Next day, the conscious rats were exposed to immobilization stress, and left ventriculography was performed (stress). In OVX rats, percentage contraction in left ventriculography was significantly reduced in response to stress, while it was not significantly changed in OVX + E rats. In both groups, heart rate was significantly increased in response to stress. However, heart rate in stress was significantly higher in OVX than in OVX + E rats. In summary, these data suggest that increase of serum estradiol levels can diminish the pathological changes in the heart induced by emotional stress.

Emotional stress triggers takotsubo cardiomyopathy in postmenopausal women. Clinical analysis of autonomic nervous function has revealed a transient increase of sympathetic nervous activity and decrease of vagal nervous activity. Immobilization (IMO) stress of rats can reproduce the electrocardiographic and left ventriculographic changes that occur in takotsubo cardiomyopathy, both of which are prevented by combined blockade of alpha- and beta-adrenoceptors. Estrogen supplementation partially attenuated these cardiac changes. It also attenuated the IMO-induced increase of c-Fos immunoreactivity, or c-fos mRNA expression in the lateral septum, medial amygdaloid nucleus, paraventricular hypothalamic nucleus, dorsomedial hypothalamic nucleus, laterodorsal tegmental nucleus, and locus ceruleus; these regions contain central sympathetic neurons and neurons with immunoreactive estrogen receptors. It also downregulated c-fos mRNA expression in the adrenal gland and the heart, suggesting an increase of estrogen attenuated the stress-induced hypothalamo-sympathoadrenal outflow from the central nervous system to the target organs. Estrogen treatment also upregulated the levels of cardioprotective substances, such as atrial natriuretic peptide and heat shock protein 70, in the heart. These data suggest that reduction of estrogen levels following menopause might be involved in the primary cause of takotsubo cardiomyopathy both by indirect action on the nervous system and by direct action on the heart.

he incidence of cardiovascular diseases is low in pre-menopausal women, whereas it is increased in post-menopausal women. Estrogen supplementation therapy prevented the increase of cardiovascular diseases in post-menopausal women, 1 while recent metaanalysis of randomized trials showed no significant merits. 2 Effects of estrogen on cardiovascular diseases were attributed principally to the modification of serum lipid concentration and coagulation pathways, while direct actions of estrogen on the cardiovascular system contributed substantially to the cardiovascular protective effects of estrogen because estrogen receptors (ER and ER ) are expressed in the blood vessels and in the heart. 3,4 Both ER and ER are also widely expressed in the central nervous system. 5 Estrogen plays crucial roles in sexual behavior, learning and memory processes, protection against ischemic insults and modulation of autonomic nervous function. 6 In fact, a reduction of estrogen levels following menopause might increase the vulnerability of women to stress while estrogen supplementation attenuates the exaggerated response to stress or to increased sympathoadrenal activity. 7 A unique form of acute cardiac attack called "takotsubo cardiomyopathy", or "transient left ventricular apical ballooning" similarly occurs predominantly in postmenopausal women in association with emotional or physical stress. [8][9][10][11][12][13][14] Although the etiology of this syndrome is yet to be clarified, an increase of serum norepinephrine, epinephrine and neuropeptide Y levels at the onset of takotsubo cardiomyopathy compared with acute myocardial infarction suggests that the exaggerated sympathoadrenal activation triggered by stress is the primary cause of this cardiomyopathy. 9,10 Immobilization stress (IMO) in the rat provides an excellent animal model of emotional stress, which activates the hypothalamic-pituitary-adrenocortical system and the sympathoadrenal system. 15 We have succeeded in developing a model of this clinical condition in rats. [16][17][18][19][20] The characteristic changes such as elevation of Circ J 2007; 71: 565 -573 (Received September 19, 2006; revised manuscript received January 9, 2007; accepted January 23, 2007 Background Takotsubo cardiomyopathy is triggered by emotional or physical stress especially in postmenopausal women. A reduction in estrogen levels following menopause might underlie the high incidence of takotsubo cardiomyopathy. Methods and ResultsThe left ventricular contraction between ovariectomized rats (OVX) and OVX with estrogen supplementation (OVX + E) while subjected to immobilization stress (IMO) was compared. The IMO in combination with general anesthesia impaired the left ventricular contraction in both OVX and OVX + E. Estrogen supplementation tended to improve the IMO-induced cardiac dysfunction and significantly attenuated the increase of blood pressure and heart rate. To understand the protective mechanism of estrogen, the expression of c-fos mRNA, a marker of cellular activation was compared. ...

In the present article the fact is emphasized that neuropeptides often are located in the same neurons as classical transmitters such as acetylcholine, 5-hydroxy-tryptamine, catecholamines, gamma-aminobutyric acid (GABA) etc. This raises the possibility that neurons produce, store and release more than one messenger molecule. The exact functional role of such coexisting peptides is often difficult to evaluate, especially in the central nervous system. In the periphery some studies indicate apparently meaningful interactions of different types with the classical transmitter, but other types of actions including trophic effects have been observed. More recently it has been shown that some neurons contain more than one classical transmitter, e.g. 5-HT plus GABA, further underlining the view that transfer of information across synapses may be more complex than perhaps hitherto assumed.

Using a paraformaldehyde-picric acid-glutaraldehyde-containing fixative and treatment of the tissue with sodium borohydride, numerous and widespread TRH-immunoreactive cell bodies were observed in the central nervous system of colchicine-treated rats, including the olfactory bulb, cortical and hippocampal areas, the caudate nucleus and other subcortical areas, many hypothalamic nuclei, the periaqueductal central gray, pontine nuclei, medulla oblongata and the dorsal horn of the spinal cord. Most of these cells could not be visualized with the same antiserum when conventional fixation methods based on formalin alone were used. The present findings suggest that TRH systems are considerably more extensive than hitherto assumed.

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Since Rinehart and Farquhar reported the presence of agranulated cells in the anterior pituitary gland in 1953, the functions of the folliculo-stellate cell remain to be clarified. Intercellular junctions have been described in the monkey, rat, and teleost anterior pituitary glands, indicating the existence of cell-to-cell communication within the organ. We pointed to their possible role in the rapid dissemination of information through a complex interconnecting system of follicles involving gap junctions. The gap junctional/folliculo-stellate cellular network was essential in the maturation and regulation of the pituitary gland system such as the hypothalamic-pituitary-gonadal axis. It has been was shown that a network participated in the conduction of electrophysiological information over a long distance using the ion Ca ϩϩ , which propagates to other folliculo-stellate cells by signaling through gap junctions. Sixty-day-old male rats were used in this study for light microscopic immunohistochemistry of S-100 protein, type I collagen, and connexin 43, and for electron microscopy to observe the morphological relationships between the cellular networks of folliculo-stellate cells and granulated pituitary cells. Clusters of anti-S-100 protein-positive cells were clearly observed in a region of the hypophysis tentatively named the transition zone. Anti-S-100 protein-positive cells and their cytoplasmic processes were also present in the anterior lobe and assembled together to form follicular lumina. Type I collagen was clearly shown outlining the incomplete lobular or ductule-like structure making cell cords in the anterior pituitary gland. Numerous microvilli were present within the follicular lumen while around the lumina, junctional specializations including gap junctions were positive for the connexin 43 protein. A nonuniform distribution of the connexin 43-positive sites were observed. Small or dot-shaped positive sites were noted where two clusters of cells were connected; the cells were identified as S-100 cells. Double immunohistochemical staining of the connexin 43 and growth hormone (GH) or connexin 43 and luteinizing hormone (LH) was also performed, demonstrating no direct relationship between the connexin 43 and either the GH or LH cells. These findings indicate that there are two kinds of messages necessary for the hormone release in the pituitary gland. One is via the portal vein system, the other is through the gap junction-mediated networks of folliculo-stellate cells. The granulated cells directly associate with cell membrane of folliculo-stellate cells are able to discharge secretory granules through communication via gap junctions, while those granulated cells that are more distant from the folliculo-stellate cells are only able to discharge hormones via the pituitary hormone-releasing hormone from the portal vein system. Anat Rec Part A 278A: 462-473, 2004.