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Several polypeptides, including Substance P, bradykinin, physalaemin and angiotensin, have been applied iontophoretically onto single neurons in the cerebral cortex of rats. The polypeptides excited many of the spontaneously active neurons tested, including identified Betz cells. Substance P was the most potent excitant, bradykinin and physalaemin were somewhat less potent, and angiotensin had only a weak excitant action. Depressant effects were observed on a few neurons. Atropine antagonized the excitant effects of acetylcholine but not those of Substance P or bradykinin. These results, taken in conjunction with other findings on the presence of some of these compounds in brain, suggest that the polypeptides deserve further consideration as possible excitatory synaptic transmitters.

The phylogenetic approach in studying hormonal actions on the kidney is important for the following reasons among others: 1) phylogenetic study of renal function has proved illuminating in the past; 2) nephron structures and functions are simpler in some non-mammalian vertebrates, and it may be easier to analyze hormonal actions on the kidney in these animals; 3) endocrine systems are different among various vertebrates, and their roles may be elucidated through comparison; 4) evolution of hormonal function has occurred, and to pursue it may be useful in determining the character of hormonal action, especially in higher vertebrates; and 5) vertebrates live in a wide variety of external environments, while the internal environment is usually kept in a narrow range. Since the kidney is a key organ for homeostasis of the internal environment, comparison of hormonal regulations of renal functions under various conditions may be instructive.

It has been established that naturally occurring pressor agents (adrenaline vasopressin and angiotensin ii) are all potent renal vasoconstrictors and, of these, vasopressin is the longer lasting and provides a back-ground of renal vasotonicity without which the kidney is unable to concentrate. The vasoconstriction affects particularly the renal outer cortical vessels and vasa recta and is associated with temporary oliguria or anuria (antidiuresis of varying degree). In association with renal outer cortical vasoconstriction there is active dilatation of the vessels supplying the adrenal gland and ovary as revealed by arteriography. Since vasopressin exerts a variable degree of renal vasotonicity according to circumstances, there appears to be an integrated feed-back system linking the neurohypophys's and hormonal responses by the adrenal and ovary through which tubular function is controlled. Since the vascular conduits between the renal artery and the adrenal gland and ovary reveal individual variation, the feed-back system induces responses which vary from modest to hyper-reactive which goes some way to explain the nature of individual responses. Taken in conjunction with other data, the evidence, strongly indicates that hormonal responses by the adrenal gland and the ovary are associated with increased blood flow. The integrated feed-back system assures that increased blood flow to the adrenal gland accompanies the appropriate signal for hormonal release. The widespread involvement of vasopressin in the systems of the body and particularly its influence on tubular function, it has been concluded, derives mainly from its vasopressor effect. The presented evidence suggests that the medullary arteries of Flint represent the original blood supply of the chromaffin mass which migrates within the adrenal cortex during foetal life. The medullary arteries and venae comites constitute an independent vascular system within the adrenal gland.

Adenylate cyclase activity and its hormonal control was measured in glomeruli isolated from collagenase treated rabbit kidneys. Experiments were performed on glycerol treated frozen glomeruli since the freezing procedure was shown to greatly enhance adenyl-cyclase activity. It was found that cAMP generation was linear 1) as a function of time up to 60 min and 2) as a function of the number of glomeruli up to 50 per sample. All further determinations were made using samples of ten glomeruli each, incubated for 30 min at 30 °C. Under such conditions, the mean control adenylate cyclase activity was found to be equal to 0.65 ± 0.04 SEM pmoles/10 gl/30 min. Glomerular adenylate cyclase was stimulated by PTH. From the mean dose response curve, an apparent k m value (2.3 × 10 −7 M) and a maximal stimulation ratio ( V max equal to 272% of the control) were calculated. PTH stimulation up to 5-fold could be observed in the most sensitive preparations. Low concentrations of arginine vasopressin produced a small but significant stimulatory effect with a mean value equal to 139 per cent of that measured under the control conditions. No effect on adenylate cyclase activity was obtained either with angiotensin II or isoproterenol.

(N-t-Butoxycarbonyl-S-acetamido-methyl-L-cysteinyl)-[1- asparagine, 5-valine]-angiotensin II (2) was prepared from [1-asparagine, 5-valine]‥angiotensin II (1). With respect to enhancement of rat blood pressure, it is a full agonist displaying about 20–25% of the potency of 1. The sulfur atom of the cysteine group represents a specific nucleophile by which the hormone can easily be attached to other molecules in a well-defined manner. Thus, 2 has been added to N-substituted maleimides through its (deprotected) thiol group to produce the spin-labelled derivative 3:{N-t-butoxy-carbonyl-S-[N-(1-oxy-2,2,5,5-tetramethyl- pyrrolidin-3-yl-methyl)-imidosuccin-3-yl]-L-cysteinyl}-[1-asparagine,5-valine]-angiotensin II, and the polymer-supported derivative 6: S-(N-{5-[7-(sepharosyl-oxy-carbonimidoyl-amino)-4-azaheptylcarbamoyl]-pentyl}-imidosuccin-3-yl)-L-cysteinyl-[1-asparagine, 5-valine]-angiotensin II. The ESR. spectra of 3 in different solvents, and its principal NMR. characteristics, are reported. We are using the compound for studying its interaction with receptor molecules and for purification of target cells and membrane vesicles by affinity aggregation.

A technique of radioimmunoassay is presented that eliminates pipetting and centrifugation, and excludes interferences by high-molecular-weight materials from the incubation and separation steps. A solid-phase binding reagent is prepared by first entrapping antibody in polyacrylamide gel. This gel is then fragmented, sieved, dried with ethanol or lyophilized, and placed in miniature disposable chromatographic columns. Application of the sample to the intra-gel column compartment is determined by the water regain of the gel. This pipetless method of sample application depends on reproducible aliquots of dry gel particles in every column. A method for preloading radiolabeled hormone and standard hormone into the column is also described. This technique has been successfully applied to the assay of angiotensin I and insulin. Dry antibody—gel stored at room temperature for 26 months has not shown loss of binding activity.

Aspartic acid was replaced in position 1 of angiotensin II (ATII) with several amino acids, to assess the possible influence of the N-terminal amino acid for (a) the intrinsic activity, (b) the affinity, and (c) the metabolic degradation of agonist analogues of ATII. Some of the substitutions in position 1 were used in combination with replacement of Phe by Gly or Leu in position 8, to obtain the corresponding antagonist.The compounds were tested in vivo (rat blood pressure) and in two in vitro preparations (rat stomach and rabbit aorta strips). The oil immersion technique, described by Kalsner and Nickerson (1968) (Can. J. Physiol. Pharmacol. 46, 719–730), was used to study the disposition of the peptides by vascular smooth muscles (rabbit aorta strips). Degradation of the peptides by purified aminopeptidases was evaluated in vitro by measuring the fragments on paper chromatography. Potency of antagonists was estimated in vivo (ID50) and in vitro (pA2 values): duration of action was established by infusing the inhibitors intravenously into anesthetized rats and testing the effect of standard doses of angiotensin before and after.The results indicate that replacement of Asp with other amino acids does not influence the intrinsic activity, but can either increase or decrease the affinity in vitro or the potency in vivo. 1-Sar-ATII, and 1-D-Ala-ATII are more potent and longer acting than 1-Asp-ATII on isolated intestinal and vascular smooth muscles, but not in vivo. On the contrary, 1-β-Asp-ATII and 1-β-D-Asp-ATII are more potent than 1-Asp-ATIIin vivo, but not on rabbit aorta strips. Rate of relaxation of rabbit aorta strips suspended in oil, after contraction with submaximal doses of several analogues of ATII, are significantly slower than relaxation after 1-Asn2-ATII and 1-Asp-ATII. A close parallelism between the diminution of the relaxation rate in oil and the degradation by aminopeptideses in vitro was observed, suggesting that metabolic degradation may be the major factor determining relaxation of rabbit aorta in oil after contraction with one of these peptides. Potencies of antagonists in vivo and in vitro are increased by replacing Asp with Sar. Substitution of Asp with β-Asp or β-D-Asp brings about a slight increase of potency in vivo but not in vitro. It appears that firm binding and prolonged occupation of receptors by sarcosyl derivatives are the primary factors contributing to increase the potency and to prolong the duration of action of antagonists, while prevention or reduction of metabolic breakdown by aminopeptidases is much less efficient.

1 The effect of calcium on the inhibition of renin secretion by biologically active angiotensin was investigated in the isolated rat kidney perfused with Krebs-Ringer saline. 2 In the presence of calcium (3.7 mM), asp(NH2)′-angiotensin II suppressed both basal and isoprenaline-stimulated renin secretion. Renal perfusion pressure, which was increased by the infusion of angiotensin, returned to control levels when isoprenaline was added. 3 When the calcium concentration was reduced to 0.32 mM, the vasoconstriction produced by angiotensin was abolished although the inhibitory effect on renin secretion was still evident. 4 In the absence of calcium, angiotensin no longer suppressed basal renin secretion and a prompt increase in renin secretion occurred when isoprenaline was added. 5 The higher basal renin levels which were observed in calcium-free perfusions, suggest the existence of an intrarenal calcium-dependent mechanism that regulates basal renin secretion. 6 These observations indicate that the inhibitory effect of biologically active angiotensin, on basal and isoprenaline-stimulated renin secretion, is functionally related to the contractor response by its dependence on calcium. The recognition that the renin-producing cells are modified smooth muscle cells supports this association

The influence of bradykinin-binding antibodies on the hypotensive and the oxytocic responses to bradykinin was investigated. Bradykinin-binding antibodies were obtained from rabbits injected with bradykinin coupled to ovalbumin. Pre- and post-immunization sera were tested for their capacity to bind intrinsically labeled 14C-bradykinin. Six injected rabbits produced bradykinin-binding antibodies. The binding ofa radio-iodotyrosine analog of bradykinin to antiserum was inhibited by unlabeled bradykinin or kallidin, but was unaffected by oxytocin, vasopressin or angiotensin. Incubation of bradykinin with antiserum resulted in an inhibition of its hypotensive activity in the rat. Bradykinin-induced hypotension was also abolished in rats injected with antiserum. Pre-immunization serum was ineffective in both respects. Bradykinin binding antibodies also inhibited the oxytocic activity of bradykinin on the isolated rat's uterus. The inhibitory effects became evident after dialysis of the antiserum. Undialyzed antisera contained an oxytocic substance which masked the inhibition of bradykinin activity. Inhibition of pharmacological effects of bradykinin by antibradykinin was reversed by heat inactivation of antibodies. Bradykinin-binding antibodies not only inhibited bradykinin activity, but also protected the peptide against kininases.

Summary The mechanism of the vasodilatory effect of furosemide on the superior mesenteric artery (SMA) was investigated in dogs. The vasodilating effect of 10 mg/kg of furosemide on the SMA was unmodified by: hydration or dehydration; bilateral nephrectomy; depletion of serotonin and catecholamines by reserpine pretreatment; methysergide blockage of serotonin; α and β adrenergic blockade, and angiotensin inhibition. It is concluded that furosemide does not cause SMA vasodilatation by inhibiting the vasoconstrictor action of serotonin catecholamines or angiotensin, and its action is probably direct on vascular smooth muscle.

The action of angiotensin on the isolated guinea-pig aortic strip is shown in relation to other agonists. The apparent dissociation constant for angiotensin, as calculated from the dose–response curve, is 6.3 × 10−8 M. The blocking action of 8-Leu-angiotensin II on this preparation is specific for angiotensin since responses to other agonists are not modified. Complete reversibility of the blockade is demonstrated. A parallel shift to the right of the dose–response curve and attainment of maximum response indicate that the antagonism is competitive for angiotensin. It is concluded that 8-Leu-angiotensin II is a highly potent antagonist for angiotensin in this tissue.

SUMMARY 1. Intracerebroventricular (icv) administration of the renin inhibitor pepstatin inhibited the dipsogenic response to icv renin but did not affect drinking induced by icv angiotensin I and II. 2. Intracerebroventricular administration of an angiotensin converting enzyme inhibitor, SQ 20881, reduced the dipsogenic effect of icv renin and angiotensin I but did not affect angiotensin II induced drinking. 3. Intracerebroventricular administration of SQ 20881 reduced the dipsogenic effect of intravenous angiotensin I but did not reduce drinking elicited by intravenous angiotensin II. 4. Intracerebroventricular administration of Sar1-Ala8-angiotensin II, a competitive antagonist of angiotensin II, inhibited drinking elicited by icv renin, angiotensin I and angiotensin II. The dipsogenic response to intravenous infusion of both angiotensins I and II was reduced after central administration of this antagonist. 5. Four analogue antagonists of angiotensin II given icv reversibly inhibited the dipsogenic response to icv angiotensin II. These analogues themselves possessed weak dipsogenic effects. 6. Renin-angiotensin induced drinking in the cat is mediated by the action of angiotensin II on receptors in the central nervous system.

During aseptic operations, dogs were fitted eiher with inflatable rubber cuffs around the descending aorta or with a polethylene tube in the cisterna cerebellomedullaris. Several weeks later they were treated with the β-Adrenoceptor-blocking agent, doberol, 5 mg/kg, and blood pressure (upstrem from aortic cuff) and heart rate were measured. The animals were anesthetized with pentobarbital or were conscious. Blood pressure increase was evoked either by inflation of the aortic cuff or by i.v. injection of 0.1–0.2 μg/kg angiotensin. In all these experiments the resulting reflex bradycardia was facilitated, when clonidine was injected either i.v. 30 μg/kg, or intracisternally (i.ci.). The α-adrenoreceptor-blocking drug phentolamine, 0.5 mg/kg, injected i.ci. antagonized this facilitatory effect of clonidine. The adrenergic nature of central neurones which stimulated the reaction pattern of ‘cardiovascular depression’ is discussed.

Melanocyte stimulating hormone release-inhibiting factor (MIF), thyrotropin releasing hormone (TRH) and angiotensin II, injected i.p. to mice, potentiate the behavioral effect of L-DOPA and 5-hydroxytryptophan.

The effect of renin depletion on autoregulation of renal blood flow and glomerular filtration rate was examined in an isolated perfused kidney preparation. Group 1 dogs received a normal-sodium diet, group 2 dogs received deoxycorticosterone acetate (DOCA) and a high-sodium diet, group 3 dogs received DOCA and a sodium-deficient diet, group 4 dogs received a high-sodium diet without DOCA, and dogs in groups 5 and 6 were treated just like dogs in groups 1 and 2, respectively. Renin release was stimulated by decreasing renal arterial pressure to 50 mm Hg. An increase in renal arterial pressure from 100 to 160 mm Hg was associated with an impaired autoregulatory response in kidneys from group 2 dogs and in two kidneys from group 4 dogs; these kidneys exhibited suppressed or absent renin release in response to a decrease in renal arterial pressure. A separate group of experiments demonstrated that DOCA plus a high-sodium diet completely depleted the kidney of renin stores, indicating that decreased renin secretion reflected depletion of renal renin content. Infusing angiotensin II (0.22 ± 0.06 µg/min) into the renal artery of group 2 kidneys did not normalize the autoregulatory response. A decrease in renal arterial pressure from 150 to 50 mm Hg was associated with impaired autoregulation of glomerular filtration rate and renal blood flow in kidneys from group 6 dogs; renin secretion was undetectable in these kidneys. All kidneys from groups 1, 3, and 5 and the remaining six kidneys from group 4 exhibited a normal autoregulatory response and normal renin release. In groups 1-3, zonal blood flow was measured using the radiolabeled microsphere technique. Only in group 2 did a significant redistribution of fractional blood flow from the inner to the outer cortex occur. These experiments demonstrate that renin depletion impairs the capacity of the kidney to autoregulate blood flow and glomerular filtration rate; thus the data are consistent with the hypothesis that the renin-angiotensin system participates in the autoregulatory response.

The pressor effect of three doses of angiotensin II (AT) and of single doses of norepinephrine (N) and tyramine (T) was measured in sodium replete and deplete rats before and after ganglionic blockade. In sodium deplete rats, plasma renin concentration (PRC) and hematocrit were significantly higher and plasma sodium was significantly lower than in sodium replete rats. The pressor effect of AT, but not that of N and of T was blunted in sodium deplete rats before and after ganglionic blockade. The relationship between the pressor effect of AT and PRC was significantly negative in the combined material of all groups before ganglionic blockade. The correlation was more pronounced after ganglionic blockade, and it became significant also in the sodium deplete group alone. The pressor effect of AT was not significantly related to plasma sodium concentration and hematocrit, while both exhibited a significant correlation with PRC. Results suggest that the influence of sodium depletion on the pressor action of angiotensin may be mediated by preexisting renin or angiotensin levels and that this relationship is influenced by the autonomous nervous system.

Ventricular weight in spontaneously hypertensive rats (F26 generation, Okamoto-Aoki strain) was significantly higher ( P < 0.001) than that in body weight-matched American Wistar and Kyoto-Wistar normotensive rats, not only among older groups of rats but also among younger groups that had not developed significant hypertension. Deoxyribonucleic acid (DNA) concentration in ventricular muscle was not different from normal in the youngest group ( P < 0.4) but was significantly reduced in the older spontaneously hypertensive rats ( P < 0.01). Plasma renin activity was significantly increased in younger spontaneously hypertensive rats before the development of established hypertension; moreover, ventricular weight and plasma renin activity were significantly correlated in younger rats ( r = 0.788, P < 0.005 for all rats, r = 0.644, P < 0.01 for spontaneously hypertensive rats). Antihypertensive therapy with either α-methyldopa or hydralazine reduced blood pressure, especially in hypertensive rats; however, ventricular weight was reduced by methyldopa ( P < 0.01) but not by hydralazine. Plasma renin activity was reduced by methyldopa but increased by hydralazine ( P < 0.01). DNA concentration was reversed toward normal by methyldopa but not by hydralazine. Similar results were obtained when methyldopa and hydralazine were given to younger rats to prevent hypertension. The changes in ventricular weight with the onset of hypertension and with its reversal or its prevention suggest that blood pressure might not be the sole factor contributing to cardiac hypertrophy in the spontaneously hypertensive rat and that the renin-angiotensin system might play a permissive role enhancing myocardial hypertrophy.

Zona glomerulosa cells were isolated from the adult rat adrenal cortex and maintained in primary monolayer culture. The morphological and functional responses of the cells to different levels of potassium in the culture medium, and to serotonin, angiotensin II, ACTH and dibutyryl cyclic AMP were investigated. Medium containing 11 mM K* stimulated aldosterone biosynthesis during the first week of culture to levels up to 100 times greater than those observed with 6 mM K*,althoug h in both cases aldosterone production eventually declined to undetectable levels. Serotonin at 0.1 and 1.0 mM initially stimulated aldosterone biosynthesis, but to lower levels than those obtained with 11 mM K*. Angiotensi n II at 10 and 100 μg/ml did not stimulate aldosterone biosynthesis by cultured rat zona glomerulosa cells. ACTH at 100 mU/ml, and dibutyryl cyclic AMP at 0.5 mM, initially stimulated aldosterone biosynthesis but later depressed i t to below levels seen in their absence. Thus none of these trophic stimuli were fo...

The effect of CRD-401 on several types of experimentally induced arrhythmias in guinea-pigs and dogs and the antagonistic effect against isoproterenol in the isolated atrium of guinea-pig were studied and compared with quinidine and propranolol. In epinephrine-induced arrhythmias of guinea-pigs, propranolol had a notable but brief depressive effect, which was observed only 5 min after administration of the drug. Although the potency of CRD-401 was weaker than that of propranolol in these arrhythmias, the effect persisted for more than 35 min after administration. In ouabain-induced arrhythmias of guinea-pigs, CRD-401 and propranolol showed similar depressive effects. Quinidine had little effect on both epinephrine- and ouabain-induced arrhythmias. Both CRD-401 and propranolol failed to reverse the arrhythmias produced by two-stage coronary artery ligation. Propranolol showed a competitive antagonism against the positive chronotropic effect of isoproterenol, while CRD-401 revealed a non-competitive β-blocking action. Quinidine produced no blocking effect in the present experiments.

A method for the isolation of guinea pig ileum smooth muscle cell membranes is described. The plasma membrane fraction possessed a (Na + , K + )-ATPase which was inhibitied by ouabain. The Mg 2+ -dependent ATPase of the membrane fraction was stimulated by 1 μM Ca 2+ . A basal ATPase, not dependent on Mg 2+ , was directly stimulated by Ca 2+ in the range of 1 μM to 1 mM. The isolated membranes contracted in response to the following substances: ATP, angiotensin II and some of its analogs, bradykinin, acetylcholine and histamine. The contractility was inhibited by ouabain and chlorambucil-angiotensin II, but not by cytochalasin B. No contraction was produced by AMP, angiotensin I and adrenaline.

The present study was undertaken to evaluate the effects of several prostaglandins (PG) on the in vitro velocity of the reaction between human renin and homologous renin substrate. In the presence of high renin concentrations, 1.4 × 10−6 moles PGA1 and PGA2 each significantly inhibited the rate of angiotensin I production (p 0.05). At lower enzyme concentrations, PGA2 was a more potent inhibitor than PGA1. In the presence of physiologic renin concentrations, 1.4 × 10−9 moles PGA2 but not 1.4 × 10−10 moles PGA2 inhibited renin reactivity. Unrelated to differences of endogenous renin substrate concentration, renin reactivity in plasma of uremic patients was greater than that of normotensive control subjects, even after the addition of 1.4 × 10−9 moles PGA (p < 0.025). Renin inhibition was demonstrated by concentrations of PGA within the renal medulla, but not at the lower concentrations in peripheral venous plasma. Consequently, if PGA induced renin inhi...

Umbilical cord plasma aldosterone consistently exceeded that of paired maternal plasma (N=24), although plasma renin activity (N=15) and reinin substrate (N=25) were always higher in the mother. This dissociation of levels of aldosterone and its stimulating hormone renin may be due to (1) a different stimulus for aldosterone secretion in the fetus, (2) increased fetal sensitivity to renin-produced angiotensin II, or (3) altered fetal metabolism of the latter hormone.

Angiotensin tachyphylaxis in the isolated guinea pig ileum and rat uterus was studied with the aid of the following synthetic peptides: [Ile5]agiotensin II (A II), [Asn1] A II, [Gly1] A II, [Arg1] A II, des-Asp1-A II, [Gly1,Gly2] A II, [Suc1] A II and acetyl-A II. The pK values for the amino and imidazole groups of these peptides were determined by electrometric titration. The effect of pH on tachyphylaxis to the angiotensin analogs and homologs was studied in the two smooth muscle preparations. The intensity of tachyphylaxis produced by the peptides correlated well with the degree of protonation of the amino group, but not with that of the imidazole group or with the net charge of the peptide. [Gly1,Gly2] A II and the two peptides in which the amino group was absent ([Suc1] A II) or blocked (acetyl-A II) did not produce any tachyphylaxis in the guinea pig ileum or the rat uterus. It is concluded that the protonated amino group plays an important role in the phenomenon of tachyphylaxis, in both organs, and that the guanido group is also important.

Angiotensin II (AII) acts as both an endogenous vasoconstrictor agent and a specific trophic hormone that controls aldosterone secretion. The characteristics of vascular and adrenal receptors for AII were examined in vitro by comparing stimulation and blockade induced by two structural analogues of AII. The adrenal response was assessed on the basis of corticosterone and aldosterone production by isolated rat glomerulosa cells; isolated rabbit aortic strips provided an index of the response of vascular smooth muscle. Threshold sensitivity to AII in both tissues was 0.3-1.0 ng/ml with a peak response at about 300 ng/ml. PII3 (I-Sar-8-Ala-AII) was a competitive antagonist in both systems; 1-10 ng/ml of PII3 produced a significant parallel shift of the All dose-response curve (P < 0.01). The maximum response was not reduced; increasing doses of AII overcame the blockade, and the responses to other agonists were not influenced. PII3 did not stimulate either tissue. P 4 T 8 (4-Phe-8-Tyr-AII) was a partial agonist in the rabbit aorta; 100 ng/ml of P 4 T 8 induced competitive blockade, and contraction followed doses above 1,000 ng/ ml. Conversely, P 4 T 8 neither stimulated nor blocked the adrenal receptors, despite very high concentrations. Thus, the results of this study reveal functional differences in the All receptors in the two systems and provide an approach to designing therapeutic agents that might block only one of its two major biological effects.

The spin-lattice relaxation times (T1) of carbon 13 in natural abundance were determined for proline, N-acetyl-prolineamide, glycylproline, cyclo(triprolyl), and a series of proline-containing peptide hormones. The data are interpreted in terms of rapid interconversion of proline between various ring-puckered forms. The nature of the puckering depends upon the type of group attached to proline. In proline, the β, γ, and δ carbon atoms are appreciably more mobile than the α carbon atom, suggesting rapid interconversion between a number of ring-puckered forms. In melanocyte-stimulating hormone release-inhibiting hormone (Pro-Leu-Gly-NH2) and its dimethylamido analog, the γ carbon atom has the greatest mobility, suggesting a rapid endo-exo interconversion at this position. In Gly-Pro and in acetyl-Pro-NH2, the cis and trans conformers of proline have very similar T1 values, indicating very little dependence of the dynamic proline ring conformation on the conformation at the peptide bond; in both isomers, the proline ring interconverts rapidly between half-chair conformers puckered at Cβ and Cγ. In thyrotropin-releasing hormone (lGlu-His-Pro-NH2), the proline ring conformation is similar to that in Gly-Pro, and independent of the state of ionization of the histidine residue. Oxytocin [see PDF for equation] , lysine-vasopressin [see PDF for equation] , [Ile5]angiotensin II (Asp-Arg-Val-Tyr-Ile-His-Pro-Phe), and luteinizing hormone-releasing hormone ( lGlu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH2) all have proline in the trans conformation about the peptide bond, and the δ carbon atom has a mobility comparable to that of the α carbon atom. The latter reflects severe steric constraints at Cδ due to the neighboring residues. In oxytocin and angiotensin II, the β carbon atom is considerably more mobile than Cα or Cδ. In luteinizing hormonereleasing hormone, the proline ring apparently undergoes rapid interconversion between half-chair forms puckered at Cβ and Cγ, whereas in lysine-vasopressin, the mobility of Cγ is greatest.

The effects of substance P and other peptides were studied on the neurons of isolated spinal cord of the frog. Substance P and certain related peptides, which have a common C-terminal sequence, -Phe-X-Gly-Leu-Met-NH2 (where X=Ile, Tyr or Phe), exerted a remarkably strong excitant action on spinal motoneurons. On a molar basis, substance P was about 200 times, physalaemin 1500 times, and eledoisin 2000 times more active than l -glutamate in depolarizing the spinal motoneuron in 0.4 mM Ca-Ringer's solution. Since the depolarizing action of substance P and related peptides persisted after the synaptic transmission was blocked by Ca-deficient (0–0.2 mM) Ringer's solution or by tetrodotoxin (10−8−10−7 g/ml), it was concluded that these peptides have a direct action on the motoneurons to induce the depolarization. Another group of peptides, bradykinin, angiotensins I and II, caused the depolarization of the spinal motoneurons by a trans-synaptic mechanism. These peptides probably activated the interneurons and thus caused the release of the excitatory transmitter from the nerve terminals synapsing with motoneurons. The possibility that substance P may be an excitatory transmitter of primary sensory neurons was discussed.

1. The adrenal response to a reduction in extracellular fluid volume (ECFV), sodium depletion, intravenous adrenocorticotrophic hormone (ACTH) and angiotensin II was assessed from changes in plasma aldosterone and plasma cortisol in eleven nephrectomized patients. Five patients were also studied before nephrectomy and six after successful renal transplantation. 2. Before nephrectomy, the resting concentrations of plasma aldosterone were usually raised. In the three patients out of five tested, the response to intravenous ACTH was marked. In two out of four patients, there was a good response to angiotensin II infusion. In one of two patients, there was a good response to a reduction in ECFV and sodium depletion. 3. After nephrectomy, the resting concentrations of plasma aldosterone were negligible or uniformly reduced and the response to ECFV reduction, sodium depletion, ACTH and angiotensin II was universally poor. 4. In all cases there was a normal plasma cortisol response to ACTH. 5. After renal transplantation in six patients, the four to whom an infusion of angiotensin II was given showed a return to a normal aldosterone response and all six showed a normal response to ACTH. 6. No clear correlation was shown between resting concentrations of plasma aldosterone and plasma potassium after nephrectomy. 7. It was concluded that the kidney, through the renin-angiotensin system, may play its main part by sensitizing the zona glomerulosa to other stimuli, but an alternative kidney-dependent mechanism cannot be excluded.

Rat adrenal capsular sections, consist-ing of the zona glomerulosa, were found to contain a substance capable of binding angiotensin II in a highly specific manner. Evidence for angiotensin II binding to a macromolecule was obtained using Sephadex gel filtration and sucrose density gradient ultracentrifugation as analytical techniques. The binding activity was temperature dependent and saturable. The binding substance was localized in particulate cell fractions and was altered by pro-teolytic enzymes. Tissue specificity studies indi-cated binding was confined almost exclusively in the zona glomerulosa of the adrenal gland. Analogues of angiotensin II known to be phar-macological agonists or antagonists effectively com-peted for angiotensin II binding sites, whereas angiotensin I, ACTH, and several other peptides were less effective. The data suggest that the binding substance in adrenal glomerulosa cells is a physiologically significant angiotensin II receptor. (Endocrinology 95: 1026, 1974)

1. Competitive or non-competitive inhibition of the myotropic and pressor response of angiotensin II is dependent on the nature of the substituent in position 8 of the antagonist peptide analogue. Substituents in other positions of the molecule, particularly position 1, contribute greatly to the potency of these antagonists. 2. As is evidenced after adrenalectomy or after blockade with phentolamine and phenoxybenzamine, the initial pressor activity observed with all the antagonistic peptides is partially due to the release of catecholamines and partially to a direct myotropic effect. 3. [Sar1, Thr8]angiotensin II has been found to possess the lowest agonist to antagonist ratio of all antagonists tested. 4. [Des-Asp1, Ile8]angiotensin II selectively and specifically inhibits the release of aldosterone from adrenal cortex. Thus, unlike angiotensin II, this heptapeptide has pronounced organ specificity, suggesting that the heptapeptide (angiotensin III) is the aldosterone-releasing hormone.