Induction by glucocorticoids of angiotensin converting enzyme production from bovine endothelial cells in culture and rat lung in vivo.

The effect of corticosteroids on angiotensin converting enzyme was investigated in endothelial cell cultures and intact rat lung. Cultured endothelial cells from bovine aorta showed net production of angiotensin converting enzyme (ACE) over 2 d culture in serum-free medium. Dexamethasone (DM) increased cell ACE activity six- to sevenfold at 100 nM with a threshold effect at 0.3 nM. The effect of DM on ACE production was completely inhibited by actinomycin D or cycloheximide. Deoxycorticosterone (DOC) and aldosterone were markedly less active, with a threshold near 100 nM and significant (two to threefold) stimulation of ACE activity at 1 muM. In cells incubated in the presence of 10 nM DM, DOC (10 muM) significantly inhibited ACE production compared with 10 nM DM alone, suggesting that DOC is a partial agonist/partial antagonist in this enzyme system. Protein content of cells or medium was unchanged by steroids at all doses used. In vivo, adrenalectomized rats showed lower pulmonary ACE compared with intact controls, and when injected with DM (40 mug/d for 4 d) showed a significant (twofold, P < 0.002) increase in lung ACE over oil-injected, adrenalectomized controls; serum ACE did not change. Injection with DOC (40 mug/d) or aldosterone (10 mug/d) had no effect on lung or serum ACE. Over a range (0.6 to 2,000 mug) of concentrations of DM administered daily for 7 d, the dose-response curve of DM for induction of pulmonary ACE mirrored that for thymolysis; for both, half-maximal effects were seen at approximately 6 mug DM/d, and plateau levels at 60 mug/d. We conclude that glucocorticoids are potent inducers of ACE activity in endothelial cells in culture and in rat lung in vivo, and that the action of aldosterone and DOC reflects occupancy of glucocorticoid receptors. This effect may be of (patho)physiological relevance in regulating levels of ACE in local vascular beds, and thereby modulating local levels of the vasoactive peptides angiotensin II and bradykinin.

Abundant calcitonin receptors in isolated rat osteoclasts. Biochemical and autoradiographic characterization.

Calcitonin receptors have been characterized for the first time in isolated osteoclasts. These receptors have been demonstrated by autoradiographic and biochemical methods, and the cells have also been shown to respond to calcitonin with a dose-dependent increase in cyclic AMP. The receptors in rat osteoclasts are specific and of high affinity (dissociation constant, Kd, 1 to 6 X 10(-10) M), and are present in greater numbers than in any cell previously studied (greater than 10(6) per cell). Chemical cross-linking of 125I-labeled salmon calcitonin to osteoclasts using disuccinimidyl suberate resulted in identification of a receptor component with a relative molecular weight of 80,000-90,000. By counting grains in autoradiographic experiments, we found that greater than 80% of specifically bound radioactivity was associated with multinucleate osteoclasts and the remainder was associated with mononuclear cells that are not osteoblasts, but that may be osteoclast precursors.

Focal induction of IGF binding proteins in proximal tubules of diabetic rat kidney.

Diabetes-associated kidney enlargement is associated with increased kidney insulinlike growth factor I (IGF-I) binding. IGF-I binds to the type I IGF receptor, which mediates most of its actions, and to specific binding proteins (IGFBPs), which modulate its actions. To explore the nature and extent of IGF-I binding in the kidney, in vitro autoradiography was used to map the distribution of IGF binding in control and diabetic rat kidney. Specificity studies were performed with increasing concentrations of unlabeled IGF-I, IGF-II, des(1-3)IGF-I (an IGF-I derivative that binds to receptors normally but with decreased affinity to binding proteins), and insulin. In control rats, diffuse binding was found throughout the kidney with increased density in the papilla. Binding specificity in the cortex and outer medulla was typical of the type I IGF receptor (IGF-I = des[1-3]IGF-I greater than IGF-II much greater than insulin). Binding in the outer medulla of diabetic kidney was typical of the type I IGF receptor. A marked focal increase in proximal tubular binding occurred in 13 of 22 postpubertal diabetic rats. Binding specificity of the proximal tubular binding was consistent with the predominance of an IGF binding protein (IGF-I = IGF-II greater than des[1-3]IGF-I with minimal displacement by insulin). Northern-blot analysis revealed increased IGFBP-1 and IGFBP-3 mRNA in cortical tissue from diabetic rats displaying increased proximal tubular binding but not from diabetic rats not displaying this phenomenon. As cell surface association of IGFBPs is linked to potentiation of IGF activity, a possible mechanism for potentiation of local IGF-I action may be provided.

The brain angiotensin system: insights from mapping its components.

Mapping of components of the angiotensin (Ang) system in the brain suggests that it serves multiple central roles, including regulation of fluid and electrolyte balance, central autonomic control, and pituitary hormone release.

The brain renin-angiotensin system: location and physiological roles.

Angiotensinogen, the precursor molecule for angiotensins I, II and III, and the enzymes renin, angiotensin-converting enzyme (ACE), and aminopeptidases A and N may all be synthesised within the brain. Angiotensin (Ang) AT(1), AT(2) and AT(4) receptors are also plentiful in the brain. AT(1) receptors are found in several brain regions, such as the hypothalamic paraventricular and supraoptic nuclei, the lamina terminalis, lateral parabrachial nucleus, ventrolateral medulla and nucleus of the solitary tract (NTS), which are known to have roles in the regulation of the cardiovascular system and/or body fluid and electrolyte balance. Immunohistochemical and neuropharmacological studies suggest that angiotensinergic neural pathways utilise Ang II and/or Ang III as a neurotransmitter or neuromodulator in the aforementioned brain regions. Angiotensinogen is synthesised predominantly in astrocytes, but the processes by which Ang II is generated or incorporated in neurons for utilisation as a neurotransmitter is unknown. Centrally administered AT(1) receptor antagonists or angiotensinogen antisense oligonucleotides inhibit sympathetic activity and reduce arterial blood pressure in certain physiological or pathophysiological conditions, as well as disrupting water drinking and sodium appetite, vasopressin secretion, sodium excretion, renin release and thermoregulation. The AT(4) receptor is identical to insulin-regulated aminopeptidase (IRAP) and plays a role in memory mechanisms. In conclusion, angiotensinergic neural pathways and angiotensin peptides are important in neural function and may have important homeostatic roles, particularly related to cardiovascular function, osmoregulation and thermoregulation.

Testicular descent. II. Ontogeny and response to denervation of calcitonin gene-related peptide receptors in neonatal rat gubernaculum.

The gubernaculum guides inguino-scrotal testicular descent by migrating into the scrotum ahead of the testis. Calcitonin gene-related peptide (CGRP) is present in the genitofemoral nerve of the neonatal rat and stimulates gubernacular motility in vitro. In a previous study in vitro autoradiography demonstrated a distinctive distribution of binding sites for CGRP over the developing cremasteric muscle in the gubernaculum. Binding analysis using computerized densitometry revealed a single class of sites. This study aimed to characterize the ontogeny of CGRP receptors in the gubernaculum and their response to denervation. Gubernacular sections from neonatal male rats were incubated with [125I]human CGRP as well as a variety of unlabeled neuropeptides. The expression of CGRP receptors culminates during the first week after birth, when gubernacular migration actually occurs. Significantly higher binding capacities were found in the denervated gubernacula compared with those in controls, which suggests an upregulation of CGRP receptors as a result of the genitofemoral nerve denervation. These results are consistent with the hypothesis that CGRP released from the nerve acts directly on the developing cremaster via its own receptors, which have not been described previously in this tissue.

Localization of bradykinin B2 receptors in the endometrium and myometrium of rat uterus and the effects of estrogen and progesterone.

In the uterus, bradykinin is a potent inducer of smooth muscle contraction, which is mediated by the bradykinin B2 receptor subtype. However, little is known about the distribution or regulation of this receptor in this tissue. The aim of this study was to localize the B2 receptor in the uterus and determine whether the levels of this receptor were altered during the estrous cycle and modulated by estrogen and/or progesterone in ovariectomized rats. At diestrus, uterine B2 receptors were localized to both the circular and longitudinal smooth muscle layers of the myometrium, the endometrial stroma, the glandular epithelium, and the layer subjacent to the luminal epithelium. B2 receptor levels in both myometrium and endometrium were lowest during early proestrus, when estrogen levels are low, whereas myometrial B2 receptor protein and messenger RNA levels were highest during late proestrous, when estrogen levels peak. Similar findings were observed for the estrogen-supplemented group after ovariectomy, with progesterone appearing to inhibit the estrogen-induced rise in bradykinin B2 receptor density in estrogen/progesterone-treated animals. Using in vitro receptor autoradiography employing the specific B2 receptor antagonist analog, HPP-HOE140, immunostaining with specific antipeptide antibodies generated against the B2 receptor, and in situ hybridization using a specific bradykinin B2 receptor riboprobe, our findings show a discrete distribution of the bradykinin B2 receptor throughout the different layers of the uterus and suggest that bradykinin B2 receptor levels in the rat uterus are regulated by estrogen, and possibly progesterone, in both myometrium and endometrium.

Localization and characterization of insulin receptors in rat brain and pituitary gland using in vitro autoradiography and computerized densitometry.

In order to identify likely sites of action in insulin in rat brain we have used the technique of in vitro autoradiography and computerized densitometry to map, characterize, and quantify its receptors in coronal and sagittal sections. A discrete and characteristic distribution of insulin receptor binding was demonstrated, with specific binding representing 92% of total binding. Displacement and specificity competition curves in olfactory bulb are typical for authentic insulin receptors, and computer analysis indicates a single class of binding site with a dissociation constant (Kd) 0.48 nM for choroid plexus and 0.44 nM for olfactory bulb external plexiform layer. Insulin receptor density is maximum in the choroid plexus, and high in the external plexiform layer of olfactory bulb. Structures of the limbic system and hypothalamus reveal moderate to high insulin receptor density, particularly the lateral septum, amygdala, subiculum, hippocampal CA1 region, mammillary body, and arcuate nucleus. Moderate insulin receptor density occurs in regions of cerebral cortex and cerebellum, and moderate to low binding occurs in discrete brainstem and midbrain structures. Insulin binding in the pituitary gland is greatest in the anterior lobe, with clear distinction from intermediate and posterior lobes. The circumventricular organs and the thalamus show low insulin binding. We conclude that insulin receptors are widespread throughout rat brain, with concentration in regions concerned with olfaction, appetite, and autonomic functions. The distribution is distinct from other neuropeptides and not related to either vascularity or cell density. A common feature of regions rich in insulin receptors is that they contain dendritic fields receiving rich synaptic input. Whether insulin plays a specific neurotransmitter or metabolic role in these sites remains unclear, but these studies have provided detailed information on potential sites of action of insulin in the brain, and will allow further studies to examine insulin receptor function in specific brain regions.

Properties of [3H]1-desamino-8-D-arginine vasopressin as a radioligand for vasopressin V2-receptors in rat kidney.

[3H]1-Desamino-8-D-arginine vasopressin [3H] DDAVP was assessed as a radioligand for vasopressin V2-receptors by studying its membrane-binding characteristics and in vitro autoradiographic localization in rat kidney, a rich source of V2-receptors. [3H]DDAVP bound specifically to a single class of high affinity, low capacity sites in rat medullopapillary membranes. Specific [3H]DDAVP binding at 25 C reached equilibrium after 2 h of incubation and was saturable and linear with protein concentration up to 2.2 mg/ml protein. Saturation analysis gave an equilibrium dissociation constant (Kd) of 0.76 nM. Displacement of [3H]DDAVP binding by unlabeled arginine vasopressin (AVP) and related analogs gave the following order of potency, consistent with that expected for a V2-receptor: DDAVP approximately equal to AVP approximately equal to 1-desamino-AVP greater than lysine vasopressin greater than oxytocin greater than [1-(beta-mercapto-beta, beta-cyclopentamethylene-propionic acid, 2-(O-methyl)tyrosine]AVP. The C-terminal metabolites of AVP, (pGlu4Cyt6)AVP-(4-9), and (pGlu4Cyt6)AVP-(4-8) did not displace [3H]DDAVP binding. No degradation of [3H] DDAVP during incubation could be detected by HPLC analysis. In vitro autoradiography of [3H]DDAVP binding to rat kidney sections showed a very dense localization of displaceable binding over inner and outer medulla, with a much lower density in cortex, consistent with the known major localization of V2-receptors on renal collecting tubules. These studies suggest that [3H]DDAVP is a suitable radioligand for labeling V2-receptors and may be useful in the characterization of vasopressin receptor subtypes in a variety of tissues and in purification of the V2-receptor.

Roles of AT1 and AT2 receptors in the hypertensive Ren-2 gene transgenic rat kidney.

Adult Ren-2 gene transgenic rats, TGR(mRen-2)27, exhibit elevated circulating and kidney angiotensin II (Ang II) levels in the presence of severe hypertension. The aim of this study was to examine whether AT1 and AT2 receptors in the kidney and renal hemodynamic and tubular responses to blockade of these receptors were altered in the Ren-2 gene transgenic rats during the maintenance phase of hypertension. Renal AT1 and AT2 receptors were mapped by in vitro autoradiography (n=8), and the effects of blockade of these receptors on mean arterial pressure (MAP), heart rate (HR), and renal cortical (CBF) and medullary blood flows (MBF) were studied in anaesthetized, adult age-matched male homozygous TGR rats (n=12) and Sprague-Dawley (SD) rats (n=7). TGR rats showed higher basal MAP (P<0.001), heart and kidney weight (P<0.001), plasma renin activity (P<0.05) and plasma Ang II level (P<0.05), and CBF (P<0.05) and MBF (P<0.05) than SD rats. AT1 receptor binding was significantly increased in the glomeruli, proximal tubules, and the inner stripe of the outer medulla of TGR rats (P<0.01), while the AT2 receptor binding was low at all renal sites of TGR and SD rats. Immunohistochemistry revealed that this increased AT1 receptor labeling occurred mainly in vascular smooth muscle layer of intrarenal blood vessels including afferent and efferent arterioles, juxtaglomerular apparatus, glomerular mesangial cells, proximal tubular cells, and renomedullary interstitial cells (RMICs) in the transgenic rats. Blockade of AT1 receptors with losartan in TGR rats markedly reduced MAP to the normotensive level (P<0.001) without altering HR. Both CBF (P<0.005) and MBF (P<0.05) were significantly increased by losartan in the transgenic rats. By contrast, losartan only caused a smaller decrease in MAP and an increase in renal CBF in SD rats (P<0.05). PD 123319 was without any renal effect in both SD and TGR rats. These findings suggest that markedly increased AT1 receptors in renal vasculature, glomerular mesangial cells, and RMICs in the presence of fulminant hypertension and elevated circulating and tissue Ang II levels may play an important role in the maintenance of hypertension in the Ren-2 gene transgenic rats.

Hypotensive action of captopril and saralasin in intact and anephric spontaneously hypertensive rats.

Intravenous injection of the converting enzyme inhibitor SQ14,225 (captopril, 2 mg/kg) reduced the blood pressure of anesthetized, spontaneously hypertensive rats (SHR) progressively over a 3-hour period. An indistinguishable fall in blood pressure occurred in SHR that were bilaterally nephrectomized 1 hour prior to injection of the converting enzyme inhibitor. In the nephrectomized animals, plasma renin activity (PRA) had fallen to less than 30% of its initial values at the time of injection. Injection of the vehicle alone had no effect on blood pressure in either anephric or intact SHR. The converting enzyme inhibitor produced no significant change in the blood pressure of either intact or anephric normotensive Wistar-Kyoto (NT-WK) rats. Infusions of Sar1-Ala8-angiotensin II (saralasin, 10 micrograms/kg-1/min-1) similarly reduced blood pressure of both intact and anephric SHR. These results indicate that captopril and saralasin lower blood pressure in the SHR by some mechanism(s) independent of the kidneys, circulating renin, or bradykinin potentiation. It is suggested that angiotensin II, locally produced at some critical tissue site(s), is involved in the maintenance of raised blood pressure in SHR.

Blood pressure responses of conscious normotensive and spontaneously hypertensive rats to intracerebroventricular and peripheral administration of captopril.

In conscious spontaneously hypertensive rats, intraxerebroventricular injection of captopril (2 mg/kg) resulted in a rapid hypotensive response that lasted several hours. The same dose given by intracerebroventricular injection had no significant effect on blood pressure (BP) of normotensive Wistar-Kyoto (WK) rats over 7 hours. There was no significant change in BP of conscious spontaneously hpertensive rats (SHR) in response to intracerebroventricular injection of vehicle and only a transitory fall in BP in response to intravenous injection of captopril (2 mg/kg). There was no significant differences between plasma renin activity (PRA) of conscious normotensive WKY rats and the PRA of SHR. These results suggest biochemical differences between the brains of SHR and normotensive WKY control rats. These differences could involve the brain renin-angiotensin system or other neuropeptides.

In vivo occupancy of angiotensin II subtype 1 receptors in rat renal medullary interstitial cells.

Angiotensin II receptor binding sites in type 1 interstitial cells in the inner stripe of the outer medulla are readily labeled in vitro by the radioligand but not in vivo after systemic radioligand administration. In anesthetized rats, we investigated if reduced vascular delivery due to angiotensin II-induced renal vasoconstriction or, alternatively, prior occupancy of these sites by endogenous angiotensins modulates angiotensin II subtype 1 receptor binding to renal medullary interstitial cells in vivo using electron microscopic autoradiography. Using 125I-angiotensin II, administered systemically, as a radioligand, binding in control rats occurred predominantly in the glomeruli and proximal tubules, while only low binding was observed in the inner stripe of the outer medulla. Pretreatment of rats with unlabeled [Sar1,Ile8]angiotensin II or with the angiotensin II subtype 1 receptor antagonist losartan before receiving the radioligand completely abolished binding to all sites. Renal vasodilatation induced by sodium nitroprusside or use of the radiolabeled antagonist analogue 125I-[Sar1,Ile8]angiotensin II did not alter binding to the inner stripe. In contrast, chronic salt loading or inhibition of angiotensin-converting enzyme by perindopril significantly increased binding not only to the cortical sites but also to the sites in the inner stripe of the outer medulla. Electron microscopic autoradiographs of the inner stripe detected binding in the interstitial cells only in rats treated with chronic salt loading or perindopril. These results suggest that endogenous angiotensins may modulate binding of circulating angiotensin II to the interstitial cells in vivo, and these angiotensin II receptor-bearing cells are more likely to be more responsive to interstitial angiotensin II than to the circulating hormone.

Inhibition of tissue angiotensin converting enzyme. Quantitation by autoradiography.

Inhibition of angiotensin converting enzyme (ACE) in serum and tissues of rats was studied after administration of lisinopril, an ACE inhibitor. Tissue ACE was assessed by quantitative in vitro autoradiography using the ACE inhibitor [125I]351A, as a ligand, and serum ACE was measured by a fluorimetric method. Following oral administration of lisinopril (10 mg/kg), serum ACE activity was acutely reduced but recovered gradually over 24 hours. Four hours after lisinopril administration, ACE activity was markedly inhibited in kidney (11% of control level), adrenal (8%), duodenum (8%), and lung (33%; p less than 0.05). In contrast, ACE in testis was little altered by lisinopril (96%). In brain, ACE activity was markedly reduced 4 hours after lisinopril administration in the circumventricular organs, including the subfornical organ (16-22%) and organum vasculosum of the lamina terminalis (7%; p less than 0.05). In other areas of the brain, including the choroid plexus and caudate putamen, ACE activity was unchanged. Twenty-four hours after administration, ACE activity in peripheral tissues and the circumventricular organs of the brain had only partially recovered toward control levels, as it was still below 50% of control activity levels. These results establish that lisinopril has differential effects on inhibiting ACE in different tissues and suggest that the prolonged tissue ACE inhibition after a single oral dose of lisinopril may reflect targets involved in the hypotensive action of ACE inhibitors.

Modulation of brain angiotensin-converting enzyme by dietary sodium and chronic intravenous and intracerebroventricular fusion of angiotensin II.

Angiotensin-converting enzyme (ACE) in rat brain closely resembled that in lung in its kinetics with the substrate Hip-His Leu, the inhibitors SQ 20,881 and SQ 14,225, and iun its Cl- activation profile. Modification of dietary NaCl intake was associated with marked changes in brain ACE activity. Sodium-loaded rats had lower activity of ACE in hypothalamus, striatum, and midbrain, and higher activity in spinal cord compared to controls. In sodium-restricted rats, ACE was elevated in pituitary and depressed in spinal cord. Chronic intravenous infusion of angiotensin (AII) was associated with a pattern of changes partly resembling sodium loading: ACE was depressed in hypothalamus and striatum but elevated in midbrain. After chronic intracerebroventricular infusion of AII, ACE was elevated in striatum and hippocampus, and depressed in spinal cord; a pattern of changes quite different from those associated with intravenous AII. These results show that ACE in several brain regions is sensitive to dietary sodium intake and support the hypothesis that angiotensin-containing neurons in these areas might be responsive to NaCl status of the animal. The observed changes in brain ACE do not seem to be explained in any simple manner by changes in circulating or central angiotensin II.

Evidence for plasticity of the dopaminergic system in parkinsonism.

A series of compensatory mechanisms within the dopaminergic system have been shown to maintain clinical function in the presence of dopamine loss. Experimental evidence for increased presynaptic dopamine turnover owing to increased dopamine synthesis, release, and reduced reuptake exists. Direct evidence that these mechanisms maintain extracellular dopamine levels is provided by intracerebral microdialysis techniques. Postsynaptic denervation supersensitivity clearly occurs with D2 dopamine receptors, although this is less evident with D1 receptors. Similarly, mechanisms of plasticity have been shown to be relevant in human postmortem and Positron Emission Tomographic studies of patients with Parkinson's disease. However, although presynaptic increases in dopamine turnover are well documented, postsynaptic D1 and D2 receptor changes have been more difficult to establish, mainly because of methodological difficulties. D2, but not D1, receptor increases have been documented in drug naive Parkinsonian patients with PET techniques. In transplantation of adrenal gland to striatum in animal models and patients with Parkinsonism where clinical improvement occurs, plasticity of host response may be as important as plasticity of the graft. Although some elements of the compensatory mechanism of dopamine plasticity may be deleterious, such as dyskinesias owing to dopamine receptor supersensitivity, the overall effect of delay and minimization of the clinical expression of disease is advantageous. An even greater understanding of the mechanisms involved may assist in developing future therapeutic strategies.

Mapping of angiotensin II receptor subtype heterogeneity in rat brain.

Angiotensin II (Ang II) exerts a number of central actions on fluid and electrolyte homeostasis, autonomic activity, and neuroendocrine regulation. In order to evaluate likely sites where these actions are mediated, Ang II receptor binding was localized in rat brain by in vitro autoradiography with the aid of the antagonist analogue 125I-[Sar1, Ile8]Ang II. Two subtypes of Ang II receptor have been identified using recently developed peptide and nonpeptide antagonists. In the periphery, the receptor subtypes differ in distribution, second messenger coupling, and function. Brain Ang II receptor subtypes were therefore differentiated into AT-1 (type I) and AT-2 (type II) subtypes by using unlabelled nonpeptide antagonists specific for the two Ang II subtypes. AT-1 binding was determined to be that inhibited by Dup 753 (10 microM) and AT-2 binding to be that inhibited by PD 123177 (10 microM). The reducing agent dithiothreitol (DTT) decreased binding to AT-1 receptors and enhanced binding to AT-2 receptors. Many brain structures, such as the vascular organ of the lamina terminalis, subfornical organ, median preoptic nucleus, area postrema, nucleus of the solitary tract, and dorsal motor nucleus of the vagus, which are known to be related to the central actions of Ang II, contain exclusively AT-1 Ang II receptors. By contrast, the locus coeruleus, ventral and dorsal parts of lateral septum, superior colliculus and subthalamic nucleus, many nuclei of the thalamus, and nuclei of the inferior olive contain predominantly AT-2 Ang II receptors. The detailed binding characteristics of each subtype were determined by competition studies with a series of analogues of angiotensin and antagonists. The pharmacological specificity obtained in rat superior colliculus and the nucleus of the solitary tract agreed well with published data on AT-1 and AT-2 receptors, respectively. There was a high degree of correlation between the distribution of Ang II binding sites with published data on Ang II-immunoreactive fields and on the sites of Ang II-responsive neurons. The present study also reveals pharmacological heterogeneity of brain Ang II receptors. The subtype-specific receptor mapping described here is relevant to understanding the role of angiotensin peptides in the central nervous system and newly discovered central actions of nonpeptide Ang II receptor antagonists.

In vitro autoradiographic localization of calcitonin binding sites in human medulla oblongata.

In this study we examined the distribution of calcitonin (CT) binding sites in the human medulla oblongata by in vitro autoradiography. In competition studies, the rank order of potency for calcitonins competing for 125I-salmon CT binding was salmon CT > porcine CT > human CT, which is consistent with physiologically relevant CT receptors in other systems. For the determination of CT binding in the human medulla, 20-micron cryostat sections were incubated with 125I-salmon CT in the presence or absence of 10(-6) M unlabelled salmon CT to map specific CT binding sites. Punctate binding was observed over discrete nuclei of the medulla. High binding densities were seen over subnuclei of the dorsal motor nucleus of the vagus, the nucleus of the solitary tract, the intermediate reticular zone, the gigantocellular and dorsal paragigantocellular nuclei, and the raphe obscurus nucleus. Moderate levels of binding were observed over the lateral paragigantocellular nucleus and the rostral extent of the epiolivary nucleus. The cuneate and gracile nuclei and the fiber tracts did not contain detectable binding, while the inferior olivary nucleus had moderate levels of nonspecific binding. The localization of calcitonin binding sites in the human presents similarities but also important differences to the distribution in rat and cat. The most notable difference is the extreme binding densities in the intermediate reticular zone of the human. The location of binding sites suggests involvement of calcitonin in regulation of autonomic function.

Localization of angiotensin II receptor binding in rabbit brain by in vitro autoradiography.

Binding of 125I-[Sar1,Ile8] angiotensin II (AII) to sections of brains from both wild and laboratory rabbits was determined by in vitro autoradiography. In the forebrain, specific high density binding was observed in the olfactory bulb, organum vasculosum of the lamina terminalis (OVLT), subfornical organ, median eminence, lateral septum, median preoptic nucleus and hypothalamic paraventricular, supraoptic and arcuate nuclei. In the midbrain, binding of the radioligand was observed in the interpeduncular and parabrachial nuclei, in the locus coeruleus, and ventrolateral pons. In the hind brain, there was dense binding of 125I-[Sar1,Ile8] AII to the nucleus of the solitary tract (NTS) and to both rostral and caudal parts of the reticular formation of the ventrolateral medulla oblongata. Weaker specific binding of the radioligand to the molecular layer of the cerebellum, to the nucleus of the spinal trigeminal tract, dorsal motor nucleus of the vagus, area postema, and to a band of tissue connecting the NTS to the ventrolateral medulla was also observed. Binding of the ligand to circumventricular organs such as the OVLT, subfornical organ, and median eminence suggests that these are sites in the brain of the rabbit at which blood-borne AII may exert influences on the central regulation of fluid balance and pituitary hormone secretion, although AII of neuronal origin could also act at these sites. Binding of the radioligand in several other brain regions suggests that angiotensin II of cerebral origin may be involved in a number of different aspects of brain function in the rabbit. The finding of dense binding in the NTS and ventrolateral medulla, which are involved in autonomic activity and are also sites of catecholamine-containing neurons, raises the possibility of angiotensin interaction with these neurons and involvement in autonomic function.

Localization and characterization of angiotensin II receptor binding and angiotensin converting enzyme in the human medulla oblongata.

Angiotensin II receptor and angiotensin converting enzyme distributions in the human medulla oblongata were localised by quantitative in vitro autoradiography. Angiotensin II receptors were labelled with the antagonist analogue 125I-[Sar1, Ile8] AII while angiotensin converting enzyme was labelled with 125I-351A, a derivative of the specific converting enzyme inhibitor, lisinopril. Angiotensin II receptor binding and angiotensin converting enzyme are present in high concentrations in the nucleus of the solitary tract, the dorsal motor nucleus of vagus, the rostral and caudal ventrolateral reticular nucleus, and in a band connecting the dorsal and ventral regions. In the rostral and caudal ventrolateral reticular nucleus, angiotensin II receptors are distributed in a punctate pattern that registers with neuronal cell bodies. The distribution and density of these cell bodies closely resemble those of catecholamine-containing neurones mapped by others. In view of the known interactions of angiotensin II with both central and peripheral catecholamine-containing neurons of laboratory animals, the current anatomical findings suggest similar interactions between these neuroactive compounds in the human central nervous system. The presence of angiotensin II receptors and angiotensin converting enzyme in the nucleus of the solitary tract, dorsal motor nucleus of vagus, and rostral and caudal ventrolateral reticular nucleus demonstrates sites for central angiotensin II to exert its known actions on vasopressin release and autonomic functions including blood pressure control. These data also suggest a possible interaction between angiotensin II and central catecholeminergic systems.