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J W Harding

Publications and source records attributed to J W Harding.

At least 55 records · Page 3Linked to original sources
PubMed1995

Characterization of a functional angiotensin IV receptor on coronary microvascular endothelial cells.

A new class of angiotensin receptors has recently been identified that exhibits both high specificity and affinity for the hexapeptide (3-8) fragment of angiotensin II, angiotensin IV (AngIV). Here, utilizing radioligand binding, we fully characterize AngIV binding at the AT4 receptor on cultured bovine coronary venular endothelial cells (CVEC), and report that when AngIV and bFGF are presented simultaneously an enhancement of DNA synthesis results that is significantly greater than that produced by bFGF alone. The level of DNA synthesis was determined by the incorporation of [3H]thymidine into quiescent CVEC monolayers following exposure to 10 nM AngIV and 10 ng/ml bFGF for 1, 3, 5, 7, 9, or 11 days. A significant enhancement of DNA synthesis (P < 0.01) was seen following 3, 5, 7, 9 and 11 days exposure. In addition, AngIV does not bind to bFGF or heparin, and conversely, bFGF is unable to compete for AngIV binding which suggests that this synergistic response is mediated by independent receptors for these ligands. Results of this study indicate that microvascular endothelial cells are significantly more responsive to bFGF in the presence of nanomolar concentrations of AngIV.

Amino Acid Sequence↗
PubMed1995

Autoradiographic identification of brain angiotensin IV binding sites and differential c-Fos expression following intracerebroventricular injection of angiotensin II and IV in rats.

A unique angiotensin binding site specific for the hexapeptide, angiotensin II(3-8) (AngIV), has been previously reported by our laboratory in the guinea pig brain and is presently described in the rat brain. This angiotensin receptor subtype has been termed AT4 and is prominently distributed in cerebral cortex, piriform cortex, hippocampus, habenulae, colliculi, septum, periaqueductal gray, several thalamic nuclei, the arcuate nucleus of the hypothalamus and cerebellum. In the second part of the present investigation, separate groups of rats received i.c.v. injections of angiotensin II (AngII), AngIV or artificial cerebrospinal fluid (aCSF) and were euthanized 2 h later for the purpose of evaluating for brain c-Fos expression. After i.c.v.-injected AngIV, Fos-like immunoreactivity was present in the hippocampus and piriform cortex. This immunoreactivity was unaffected by i.c.v. pretreatment with the AT1 angiotensin receptor antagonist DuP 753 (losartan) or the AT2 receptor ligand PD123177 but was blocked by the AT4 angiotensin receptor antagonist, divalanal-AngIV. I.c.v. injection of AngII resulted in Fos-like immunoreactivity in the dorsal third and lateral ventricles, subfornical organ, lateral hypothalamus and amygdala. Pretreatment with losartan or PD123177 significantly interfered with this AngII-induced immunoreactivity while divalanal-AngIV did not. These results indicate that in both guinea pig and rat brains the AT4 receptor has a distribution different than that previously reported for AT1 and AT2 receptor subtypes. The c-Fos expression results suggest that different brain neuronal pathways are activated by i.c.v. injection of AngII and AngIV.

Angiotensin II↗
PubMed1995

The angiotensin IV system: functional implications.

The brain renin-angiotensin system has been implicated in the central regulation of the cardiovascular system, body water balance, and cyclic regulation of reproductive hormones and behaviors. It also exerts some influence over the secretion of pituitary hormones. This system appears to be complete with the necessary precursors and enzymes for the formation and degradation of biologically active forms of angiotensins and several binding subtypes that are presumed to mediate these and other functions. Much information is now available on the AT1 site which preferentially binds angiotensin II (AngII), but also binds angiotensin III (AngIII), and appears to be responsible for mediating the above described classic angiotensin physiologies and behaviors. Less is known about the functional importance of the AT2 site which also binds AngII but preferentially binds AngIII. This site has been implicated in vascular growth and cerebral blood flow. Recently, an AT4 site has been discovered and characterized that preferentially binds AngII (3-8), a fragment of AngII referred to as angiotensin IV (AngIV). This AT4 site is prominent in cerebral cortex, hippocampus, basal ganglia, cerebellum, and spinal cord, as well as several peripheral tissues including kidney, bladder, heart, spleen, prostate, adrenals, and colon. The AT4 site may mediate memory acquisition and recall and the regulation of blood flow. The function(s) of the AT4 receptor subtype in peripheral tissues is currently unknown, although it does appear to be involved in kidney blood flow.

Angiotensin II↗
PubMed1995

Anomalous effects of losartan on aminopeptidase-induced reductions of blood pressure in SHR.

The present investigation initially determined that a commercially available aminopeptidase M (AmM, Sigma Chemical) can lower blood pressure when intracerebroventricularly (ICV) infused in spontaneously hypertensive rats (SHR). Pretreatment with the angiotensin II (AngII) receptor subtype 1 (AT1) antagonist, DuP 753 (losartan) significantly attenuated this hypotensive effect, in a dose-dependent manner, while pretreatment with the AngII receptor subtype 2 (AT2) antagonist, PD123177, did not influence AmM-induced hypotension. These results suggest that AT1 receptors may be involved in the hypotension accompanying the ICV infusion of AmM; however, the relationship among available AT1 sites, angiotensin ligands, and peptidase activity appears to be complicated with the likely involvement of additional, as yet unspecified, brain peptide systems possessing cardiovascular action.

Angiotensin Receptor Antagonists↗
PubMed1994

Brain angiotensin receptor subtypes in the control of physiological and behavioral responses.

This review summarizes emerging evidence that supports the notion of a separate brain renin-angiotensin system (RAS) complete with the necessary precursors and enzymes for the formation and degradation of biologically active forms of angiotensins, and several binding subtypes that may mediate their diverse functions. Of these subtypes the most is known about the AT1 site which preferentially binds angiotensin II (AII) and angiotensin III (AIII). The AT1 site appears to mediate the classic angiotensin responses concerned with body water balance and the maintenance of blood pressure. Less is known about the AT2 site which also binds AII and AIII and may play a role in vascular growth. Recently, an AT3 site was discovered in cultured neoblastoma cells, and an AT4 site which preferentially binds AII(3-8), a fragment of AII now referred to as angiotensin IV (AIV). The AT4 site has been implicated in memory acquisition and retrieval, and the regulation of blood flow. In addition to the more well-studied functions of the brain RAS, we review additional less well investigated responses including regulation of cellular function, the modulation of sensory and motor systems, long term potentiation, and stress related mechanisms. Although the receptor subtypes responsible for mediating these physiologies and behaviors have not been definitively identified research efforts are ongoing. We also suggest potential contributions by the RAS to clinically relevant syndromes such as dysfunctions in the regulation of blood flow and ischemia, changes in cognitive affect and memory in clinical depressed and Alzheimer's patients, and angiotensin's contribution to alcohol consumption.

Animals↗
PubMed1994

AT4 receptor structure-binding relationship: N-terminal-modified angiotensin IV analogues.

The effect of structural changes in the N-terminal amino acid of AIV, with respect to AT4 receptor binding, was examined by competition with [125I]AIV in bovine adrenal membranes. Analogues with modifications of the first residue alpha-amino group possessed lower affinities than the primary amine-containing parent compound. Peptides with a residue 1 alpha-carbon in the D conformation exhibited poor affinity for the AT4 receptor. Modifications of the residue 1 R-group demonstrate that a straight chain aliphatic moiety containing four carbons is optimal for receptor-ligand binding, as evidenced by the extremely high affinity of [Nle1]AIV (Ki = 3.59 +/- 0.51 pM). Replacement of the 1-2 peptide bond of AIV with the methylene bond isostere psi (CH2-NH), increased the Ki approximately fivefold, indicating that the peptide bond may be replaced while maintaining relatively high-affinity receptor binding.

Adrenal Glands↗
PubMed1994

Noise-induced elevations of plasma endothelin (ET-3).

The endothelins (ETs) are a novel family of peptides which participate in hemodynamic homeostasis. Elevated levels of circulating ETs are evident in several stress related conditions and are associated with a variety of vascular pathophysiologies. The purpose of the current study was to test the possibility that plasma concentrations of endothelin increase following noise exposure using radioimmunoassay (RIA). No difference in plasma endothelin was detected in rats subjected to brief noise exposure (30 min of 100 dB SPL broad-band noise) compared to control animals. Statistically significant elevations in plasma endothelin (ET-3) were measured in animals exposed to prolonged noise exposure (90 min and 72 h of 100 dB SPL broad-band noise). These results suggest that hemodynamic alterations, and potential vascular pathophysiologies accompanying prolonged exposure to noise are mediated by endothelin.

Acoustic Stimulation↗
PubMed1994

AT4 receptors: specificity and distribution.

The AT4 receptor specifically binds Angiotensin (Ang) IV and is distinct from the AT1 and AT2 receptors which bind Ang II and Ang III. The AT4 receptor in bovine adrenal cortex has a Kd of 0.74 +/- 0.14 nM and a Bmax of 3.82 +/- 1.12 pmol/mg prot. Competition curves demonstrated the following rank order of affinity: Ang IV >> Ang III >> d-Arg - Ang II >> Sar1,Ile8 - Ang II = Ang II = Ang II (1-7) >> DuP753 = CGP42112A = PD 123177. AT4 receptors were present in many tissues from several mammalian species including human and monkey. AT4 and AT1/AT2 receptors revealed a differential distribution in the rat kidney.

Adrenal Cortex↗
PubMed1993

Differences between perinatal angiotensin binding in the brains of SHR and WKY rats.

A growing body of evidence suggests that angiotensin may have a functional role in growth and development, in addition to its classical role in the maintenance of body water homeostasis. Components of the renin-angiotensin system have been identified in the rat fetus. Because of the association between the renin-angiotensin system and hypertension, we quantified angiotensin receptor binding sites in the brains of spontaneously hypertensive rats (SHR) and Wistar Kyoto (WKY) rats during perinatal development. Using in vitro receptor autoradiography we identified specific 125I-Sar1,Ile8 AII binding in several areas of the brains of perinatal rats of both strains and observed significant differences in the concentration of binding sites, at different ages in several brain nuclei. With the knowledge that components of the renin-angiotensin system appear early in development and are known to have an association with cellular growth, it is possible that an irregularity in this system occurring during neurogenesis could contribute to developmental abnormalities, as well as subsequent hypertension.

1-Sarcosine-8-Isoleucine Angiotensin II↗
PubMed1993

Identification and characterization of a novel angiotensin binding site in cultured vascular smooth muscle cells that is specific for the hexapeptide (3-8) fragment of angiotensin II, angiotensin IV.

This study demonstrates the existence of a previously unrecognized class of angiotensin binding sites on vascular smooth muscle that exhibit high affinity and specificity for the hexapeptide (3-8) fragment of angiotensin II (AngIV). Binding of [125I]AngIV is saturable, reversible and describes a pharmacologic profile that is distinct and separate from the classic AT1 or AT2 angiotensin receptors. Saturation binding studies utilizing cultured vascular smooth muscle cells obtained from bovine aorta (BVSM) revealed that [125I]AngIV bound to a single high affinity site with an associated Hill coefficient of 0.99 +/- 0.003, exhibiting a KD = 1.85 +/- 0.45 nM and a corresponding Bmax = 960 +/- 100 fmol mg-1 protein. Competition binding curves in BVSM demonstrated the following rank order effectiveness: AngIV > AngII(3-7) >> AngIII > Sar1,Ile8 AngII > AngII > AngII(1-7) > AngII(4-8), DuP 753, PD123177. The presence of the non-hydrolyzable GTP analog GTP gamma S, had no effect on [125I]AngIV binding affinity in BVSM. The presence of this novel angiotensin binding site on smooth muscle in high concentration suggests the possibility that this system may play an important, yet unrecognized role in vascular control.

Amino Acid Sequence↗
PubMed1993

Motion-induced aversions during and after recovery from olfactory nerve section in mice.

Bilaterally nerve-sectioned male albino mice were given access to a novel food (almond) and then body rotated for 50 min. Two days later the mice were given a second access to almond. The procedures were repeated after recovery of olfactory function. Mice formed a presumably taste-mediated aversion while anosmic and a second flavor-mediated aversion (to the same food) after the return of olfactory function. Rotation control subjects formed only the standard flavor aversion subsequent to the first pairing of novel food and motion. It appears that olfaction is required for recognition of a food as novel.

Animals↗
PubMed1993

Hypothalamic angiotensin release in response to AII or glutamic acid stimulation of the SFO in rats.

Recent evidence from our laboratory suggests that angiotensin II (AII) is synthesized, stored within cells in the paraventricular nucleus (PVN) of the hypothalamus, and upon appropriate stimulation, released and rapidly converted to angiotensin III (AIII). The present investigation extends these observations by first employing a retrograde tracer to confirm a direct connection from the subfornical organ (SFO) to the PVN, and then showing that microinfusion of AII or glutamic acid into the SFO provokes release of endogenous angiotensin within the PVN. Potentially it is this release that contributes to the elevations in blood pressure and drinking that have been reported to occur with electrical and chemical stimulation of the SFO. These results represent the first evidence of releasable angiotensin provoked by the chemical activation of a neural pathway that has been histochemically demonstrated to link the SFO with the PVN and brain stem structures concerned with cardiovascular functioning.

Angiotensin II↗
PubMed1993

Angiotensin II(3-8) (ANG IV) hippocampal binding: potential role in the facilitation of memory.

The present research characterizes a newly discovered ANG II(3-8) (ANG IV) binding site localized in structures associated with memory function (hippocampus, neocortex, cerebellum), as well as other brain stem structures (thalamus, inferior olivary nucleus). This site is not the AT1 or AT2 site that binds angiotensins II (ANG II) and III (ANG III) nor does it bind the nonpeptide AT1 or AT2 receptor antagonists DuP753 and PD123177, respectively. The intracerebroventricular (ICV) infusion of ANG IV was ineffective at inducing drinking in rats as compared with equivalent doses of ANG II and III. Although not as effective as ANG II or ANG III, ICV infusion of ANG IV did provoke a pressor response at the highest dose (100 pmol/min), which appeared to be mediated by ANG II (AT1)-type receptors and not the specific AIV binding site described here. By contrast, the ICV infusion of ANG IV resulted in greater effects upon retention and retrieval of a passive avoidance task as compared with ANG II. Specifically, ANG II was not different from the ICV infusion of artificial cerebrospinal fluid, while ANG IV improved retention and retrieval of this task.

Angiotensin II↗
PubMed1993

GABA and bicuculline-induced blood pressure changes in spontaneously hypertensive rats.

The brain GABAergic system was previously shown to influence blood pressure (BP) maintenance in rats which may in part be accomplished by disruption of the central renin-angiotensin system (RAS). We examined the potential role of GABA in sustaining the high BP exhibited by the spontaneously hypertensive rat (SHR) model of human essential hypertension. Intracerebroventricular (i.c.v.) infusion of GABA produced decreases in BP in members of three rat strains, including Wistar-Kyoto (WKY) and Sprague-Dawley normotensive controls and SHR. The SHR were significantly more sensitive to GABA than the normotensive strains. Next, the GABA receptor antagonist bicuculline (BMI) was infused i.c.v. and produced increases in BP in members of each strain. Finally, i.c.v. pretreatment with the specific angiotensin receptor antagonist [Sar1, Thr8]AII (sarthran), blocked subsequent GABA-induced decreases in BP in members of all three strains, and there was a trend toward sarthran attenuation of BMI-induced increases in BP. These results encourage the hypothesis that the hypotensive effects produced by central application of GABA are mediated by the brain angiotensin system.

Analysis of Variance↗
PubMed1993

Elucidation of a specific binding site for angiotensin II(3-8), angiotensin IV, in mammalian heart membranes.

Data are presented describing a new angiotensin binding site in rabbit and guinea pig heart, distinct from AT1 and AT2, that demonstrates high specificity and affinity for the hexapeptide fragment angiotensin II(3-8), which will be referred to here as angiotensin IV (AIV). Equilibrium binding in rabbit heart membranes was achieved in 2 hr at 37 degrees C and produced a calculated kinetic KD of .174 +/- .018 nM. Saturation equilibrium binding data for rabbit and guinea pig heart were best fit to a one-site model with Hill coefficients near unity. Guinea pig membranes exhibited a KD = 1.33 +/- .02 nM and a Bmax = 144 +/- 19 fmol/mg protein, and rabbit heart membranes had a KD = 1.70 +/- .50 nM and a Bmax = 731 +/- 163 fmol/mg protein. The binding site showed a high specificity for AIV, although it exhibited low affinity for angiotensin II, angiotensin III, Sar1,Ile8-angiotensin II, DuP 753, CGP42112A and PD123177. A large number of nonangiotensin-related peptides were unable to compete effectively for 125I-AIV binding. Deletions made from the C-terminal end of AIV caused a decrease in affinity: AIV > AII(3-7) >> AII(3-6) >> AII(3-5). Extension of the C-terminal end of AIV corresponding to the amino acids of human angiotensinogen caused little change in affinity. GTP gamma S had no effect on binding, suggesting non-G protein linkage. Binding was widely distributed throughout the heart; it was observed on cardiocytes and blood vessels as well as in the epicardium and the endocardium.

Amino Acid Sequence↗
PubMed1993

Central angiotensin IV binding sites: distribution and specificity in guinea pig brain.

Our laboratory has reported previously that a unique binding site specific for the hexapeptide angiotensin (A)II(3-8), now referred to as AIV, is present in a number of tissues including bovine adrenal gland, rabbit and guinea pig heart and guinea pig kidney, liver, lung, uterus and brain. The present results extend previous findings in the guinea pig brain and identify binding sites for AIV in the neocortex, paleocortex, hippocampus, medial habenula, superior and inferior colliculi, caudate putamen, thalamus, dorsal tegmentum, central gray, red nucleus, inferior olivary, oculomotor and hypoglossal nuclei and cerebellum. Binding of [125I]AIV in selected regions was shown to be of high affinity (Kd = 0.60-1.47 nM), saturable (maximal number of binding sites = 181-449 fmol/mg of protein) and specific. This binding site was shown to be distinct from the AT1 and AT2 sites with Ki values > 10(-4) M for DuP 753, CGP42112A and PD123177. Changes at the N-terminal of the peptide, either by removal of the valine or by extension of the peptide, resulted in a large decrease in binding affinity. In contrast, C-terminal extensions resulted in little change in affinity for the binding site. Guanosine 5'-0-(3-thiotriphosphate) was shown to have no effect on binding, suggesting that the guinea pig brain binding site is not G-protein-linked. Potential functions associated with this newly discovered A binding site are discussed.

Amino Acid Sequence↗
PubMed1992

Identification of an AII(3-8) [AIV] binding site in guinea pig hippocampus.

A unique angiotensin binding site specific for the hexapeptide, AII(3-8), has been identified in guinea pig hippocampus. This binding site, which is present in the pyramidal cell layer of CA1, CA2, CA3 of the hippocampus and dentate gyrus, binds AII(3-8) with high affinity (KD = 1.29 +/- 0.18 nM) in a saturable manner (Bmax = 449 +/- 62 fmol/mg protein). The N-terminal structure of the binding ligand is paramount in determining the binding affinity. The C-terminal requirements seem less stringent as evidenced by the binding affinity of AII(3-7) (KD = 20.9 +/- 2.1 nM). Neither AII, AIII,Sar1, Ile8-AII, Dup 753 nor CGP42112A appear to bind, indicating that this binding site is neither the AT1 nor AT2 sites described for AII/AIII. Autoradiographic analysis of hippocampus binding confirms the inability of Sar1,Ile8-AII to compete for [125I]AII(3-8) binding. Conversely AII(3-8) was unable to displace [125I]Sar1,Ile8-AII binding.

Angiotensin II↗