M. Ian Phillips

Regional study of cerebral ventricle sensitive sites to angiotensin II.

Angiotensin II injected in small doses into the cerebral ventricles produces an increase in blood pressure and drinking behavior. The site of action for both of these effects was studied in 3 main experiments. (1) The response to several doses of angiotensin delivered to each ventricle was investigated with multiple ventricular cannulation. This revealed that the rostral ventricular system was involved in angiotensin II mediated responses. (2) CSF flow was limited by plugging specific anterior and posterior ventricular regions and then testing for angiotensin II induced drinking and pressor responses. This technique showed that the ventral anterior third ventricle must be reached by the peptide in order to produce either blood pressure or drinking effects. (3) In order to separate pressor components due to vasopressin release and sympathetic activation, hypophysectomized rats were also tested. The experiment showed that the pressor response to intraventricular angiotensin II is due to both sympathetic and pituitary hormonal components and both are dependent on sites sensitive to angiotensin in the anterior third ventricule. The ventral anterior third ventricle or periventricular tissue surrounding it seems to be essential for both blood pressure and drinking responses to intraventricular angiotensin II.

The central and peripheral effects of Captopril (SQ 14225) on the arterial pressure of the spontaneously hypertensive rat

An orally active inhibitor of the enzyme that converts angiotensin I to angiotensin II (All) and degrades bradykinin has recently been shown to lower blood pressure in spontaneously hypertensive (SH) rats when administered orally, but to have no depressor effect on normotensive Wistar-Kyoto ( W K Y ) r a t s 7,1°. Since Captopril is a small molecule (2-D-methyl-3-mercapto-propranoyl-L-proline) it might be able to cross the blood-brain barrier and exert its hypotensive action in the brain. On the other hand, it could act peripherally. I f it crosses the blood-brain barrier the action might be due to the inhibition of angiotensin II formation since it has been shown that saralasin, an angiotensin II antagonist, lowers blood pressure in SH rats when injected into the brainS, 11. Therefore, to determine if Captopril has a central action we have compared administration of the same dose intracerebroventricularly (i.v.t.) and intravenously (i.v.) to SH rats. We reasoned that if the same dose of Captopril produces a greater hypotensive response i.v.t, than i.v. it must be acting centrally and not merely leaking into the blood. This was found to be the case. Six male SH rats, 4--5 months of age, weighing 300-350 g, were anesthetized with 20 ~ chloral hydrate (2 ml/kg). Stainless-steel cannulae were implanted into the left lateral ventricles and heparinized saline-filled catheters were introduced into the left femoral arteries and veins. The venous catheter was polyethelene tubing (PE 50). The arterial catheters were Silastic inside the artery. The Silastic part of the catheter was attached to polyethylene tubing (PE 50) outside the artery. Both catheters were exteriorized by placing them subcutaneously to a cut in the skirt over the back of the rat approximately at the level of the scapulae. The catheters were held in place on the back by wound clips and cranioplastic dental cement. The rats were allowed to recover for at least one day after the operation. All experiments were performed on conscious, unrestrained rats. Mean arterial

Rat brain cells in primary culture: characterization of angiotensin II binding sites.

The binding kinetics of angiotensin II (ANG II) have been studied in primary cultures from fetal rat brain. Binding of [125I]ANG II to rat brain cells in culture is time-, pH- and cell concentration-dependent. The binding is saturable, reversible, and 90--95% specific. Binding follows first-order kinetics, with values for K1 and K-1 of 4.9 x 10(6)M-1 S-1 and 3.33 x 10(4)S-1 respectively. Scatchard analysis reveals the presence of a single class of binding sites with Ka of 1.0 x 10(9)M-1 and an average of approximately 6 x 10(3) sites per cell. [125I]ANG II recovered from incubation medium under the conditions of the binding assay or after dissociation from cells is not significantly degraded as judged by gel filtration on Sephadex G-25 and radioreceptor assay. ANG II analogs compete with [125I]ANG II for binding, with potencies in general paralleling previously established biological activities. Of 5 analogs tested, (Ile8)-ANG II was almost equipotent with ANG II while (Dval3)-ANG II was least potent in the competitive binding assay. These data fulfill criteria for the identification of specific angiotensin II receptors in cells from mammalian brain.

SENSITIVE SITES IN THE BRAIN FOR THE BLOOD PRESSURE AND DRINKING RESPONSES TO ANGIOTENSIN II

Publisher Summary The integration of water balance, which is essential for terrestrial beings, is a function of the brain. Angiotensin offers the opportunity to unravel the complex circuitry involved by leading to the receptors from which neural circuits emanate into the brain that control thirst, antidiuretic hormone (ADH) release, and sympathetic activation of blood pressure. The chapter discusses that angiotensin II receptors are not scattered all over the brain but are in circumscribed versions close to the ventricles. They are available to either blood-borne angiotensin, as in the subfornical organ and area postrema, or they are available to cerebrospinal fluid (CSF)-borne angiotensin in the ventral anterior third ventricle. In view of the proximity of the paraventricular neurons to the ventricles, angiotensin in the CSF is able to act on ADH secreting cells directly. There is some neurophysiological evidence to support this view for supraoptic nuclei. Angiotensin II produces its effects by simultaneously acting on closely proximated receptors that are the internal points of origin for different neural pathways.

Insulin, Insulin-like Growth Factors, and Their Receptors in the Central Nervous System

Section I: Insulin Receptors: Structure and Function.- Internalization of Insulin and Its Receptor: Role in Signaling.- Insulin-activated Phosphorylation on Tyrosine of a 15 Kilodalton Cytosolic Protein in 3T3-L1 Adipocytes.- Role of Protein Phosphorylation in Growth Factor Signal Transduction..- The Processing and Transport of Peptide Hormones across Endothelial Cell Barriers.- The Role of Phospholipid Metabolism in Insulin Action.- Glucose Transporters: Overview and Implications for the Brain.- Alteration of Insulin Receptor Binding and Protein Kinase Activity in Rat Liver and Placenta by ?-Naphthoflavone.- Section II: Insulin and Insulin Receptors in the Central Nervous System.- Evolution of Insulin and Insulin Receptors.- Evidence for Central Nervous System Insulin Synthesis.- Localization of Insulin to Neuronal Cells.- Synthesis of Insulin or a Similar Peptide in the Pituitary Gland and in Retinal Muller Cells.- Insulin in the Central Nervous System: A Regulator of Appetite and Body Weight.- Insulin in the Brain: A Feedback Loop Involving Brain Insulin and Circumventricular Organs.- Structural Evidence for a Subtype of Insulin Receptor in the Central Nervous System.- Physiologically Unique Insulin Receptors on Neuronal Cells.- Insulin Receptors in Brain Development.- Insulin Downregulates Alpha-2-adrenergic Receptors in Cultured Glial Cells.- Section III: Insulin-Like Growth Factor and Insulin-Like Growth Factor Receptors in the Central Nervous System.- Somatomedins (Insulin-like Growth Factors) and the Nervous System.- Insulin-like Growth Factor Receptors in the Brain.- Receptors for Insulin and Insulin related Peptides in the CNS: Studies of Localization in Rat Brain.- Visualization of IGF-2 Receptors in Rat Brain.- Insulin-like Growth Factors and Their Receptors in the Pituitary and Hypothalamus.- Two Types of Receptors for Insulin-like Growth Factors Are Expressed on Normal and Malignant Cells from Mammalian Brain.- Role of Insulin, Insulin-like Growth Factors and Nerve Growth Factor in Neurite Formation.- Contributors.- Author Index.

Recent advances in cellular and molecular aspects of angiotensin receptors

1. Characterization of a cis-Regulatory Element and trans- Acting Protein That Regulates Transcription of the Angiotensin II Type 1A Receptor Gene.- 2. Human AT1 Receptor Gene Regulation.- 3. Regulation of Gene Transcription of Angiotensin II Receptor Subtypes in the Heart.- 4. Sodium Induced Regulation of Angiotensin Receptor 1A and 1B in Rat Kidney.- 5. Characterization and Regulation of Angiotensin II Receptors in Rat Adipose Tissue: Angiotensin Receptors in Adipose Tissue.- 6. Changes in Angiotensin AT1 Receptor Density during Hypertension in Fructose-Fed Rats.- 7. Cardiac Effects of AII: AT1A Receptor Signaling, Desensitization, and Internalization.- 8. AT1-Receptors and Cellular Actions of Angiotensin II in Neuronal Cultures of Stroke Prone-Spontaneously Hypertensive Rat Brain.- 9. Antisense Oligonucleotides for in Vivo Studies of Angiotensin Receptors.- 10. Interactions of Angiotensin II with Central Dopamine.- 11. Regulation of the Hypothalmic-Pituitary-Adrenal Axis and Vasopressin Secretion: Role of Angiotensin II.- 12. Relationship between the Drinking Response to Angiotensin II and Induction of fos in the Brain.- 13. Identification of AT1 Receptors on Cultured Astrocytes.- 14. Structure-Activity Relationship of the Agonist-Antagonist Transition on the Type 1 Angiotensin II Receptor the Search for Inverse Agonists.- 15. Molecular Cloning and Expression of Angiotensin II Type 2 Receptor Gene.- 16. Molecular Cloning of the Human AT2 Receptor.- 17. Molecular and Functional Characterization of Angiotensin II AT2 Receptor in Neuroblastoma N1E-115 Cells.- 18. Characterization of the AT2 Receptor on Rat Ovarian Granulosa Cells.- 19. AT2 Receptor Expression in Ovaries: A Review.- 20. Heterogeneity of Rat Angiotensin II AT2 Receptor.- 21. Heterogeneity of Angiotensin Type 2 (AT2) Receptors.- 22. Angiotensin II Stimulates Protein Phosphatase 2A Activity in Cultured Neuronal Cells via Type 2 Receptors in a Pertussis Toxin Sensitive Fashion.- 23. Functional Aspects of Angiotensin Type 2 Receptor.- 24. Angiotensin Receptor Heterogeneity in the Dorsal Medulla Oblongata as Defined by Angiotensin-(1-7).- 25. Atypical (Non-AT1, Non-AT2) Angiotensin Receptors.- 26. Brain Angiotensin II and Related Receptors: New Developments.- 27. Receptors for (3-8) Angiotensin in Brain Cells: AngIV Binding in Brain Cells.

Independent receptors for pressor and drinking responses to central injections of angiotensin II and carbachol

Angiotensin II and carbachol when injected in the brain ventricles of the rat produce similar responses of an increase in blood pressure and drinking behavior. The question of whether these effects are produced by independent receptors or via a cholinergic circuit is debatable for the drinking behavior and evidence is lacking for the blood pressure effect. We have used a chronic rat preparation for recording blood pressure and drinking at the same time during intraventricular injections (i.v.t.) of both angiotensin and carbachol and i.v.t. or intravenous infusions of appropriate antagonists. The results show that drinking and blood pressure response to angiotensin II can be blocked by P113 (500 ng 1.v.t.) an angiotensin antagonist; they are not blocked by atropine (10 mug i.v.t.) a cholinergic antagonist; carbachol effects, however, are not blocked by P113, but are totally blocked by atropine (10 mug i.v.t.), At high doses of atropine there is inhibition of both agents but this probably represents a general inhibition. The hormone and cholinomimetic administered together interact and both are inhibited by adrenergic stimulation. We conclude from these experiments that angiotensin and carbachol act upon independent receptors in the brain to produce blood pressure and drinking responses but at some point they share common, central effector pathways.

Staining of human and rat brain Vibratome sections by a new Golgi method.

Abstract A simple rapid procedure is described for processing human and animal brain tissue with a Golgi stain that preserves the tissue. Vibratome cut sections are prepared from the tissue without embedding the block. The sections are processed within one week. The method gives good resolution of dendritic spines. It requires less work time than standard procedures and none of the tissue is lost.

Attenuation of the central hypertonic NaCl pressor response by angiotensin II inhibition.

Both angiotensin II (Ang II) and hypertonic NaCI raise blood pressure, induce drinking and release vasopressin (VP) when administered into the brain ventricles (IVT) 1,2,6,11,17,z3. The striking similarity in the effects of these two stimuli has induced investigators to study the possibility that they act through a common pathway. Several studies have suggested interactions between these two stimuli. Andersson has proposed that Ang II facilitates Na + effects by action on the Na+/K + ATPase z. Central injections of saline plus angiotensin were more effective in eliciting drinking than either stimulus alone. The sensitive sites in the brain for Ang II and hypertonic NaCI appear to be very close to each other or the same site since small electrolytic lesions of the brain region near the anteroventral third ventricle (AV3V) attenuate the pressor, antidiuretic 3 and drinking1, 5 responses. The AV3V area of the brain, which includes the organum vasculosum of the lamina terminalis, appears to be one of the most sensitive receptive areas in the brain for Ang 1115. Similarly, this region has been implicated in the reception of hyperosmotic stimuli for the induction of thirst 6, antidiuresis and pressor responses 3. In addition to the evidence that suggests the two stimuli act on the same receptive region in the brain, electrophysiological studies reveal an interaction between Ang II and hyperosmotic stimuli. Wayner et al. 2B recorded from hypothalamic neurons which were sensitive to both Na + and Ang II. These cells had a greater discharge frequency to the combination than either Na + or Ang II alone. The question that these experiments have not answered, however, is whether Ang II in the brain acts through osmoreceptors or osmotic stimuli are mediated by Ang II. Direct evidence implicating Ang II as a mediator of the osmotic stimuli for the release of VP has been provided by Sladek and Joynt 25. Using explants of the hypothalamo-neurohypophyseal system, they demonstrated that VP was released by

Different times of development of tremor to harmaline and oxotremorine in neonatal rats.

Neonatal rats at 1-2 days after birth were injected with the tremorogenic drugs harmaline and oxotremorine. The onset of tremor to the drugs was significantly different (P < 0.01). Tremor induced by oxotremorine first appeared at day 4 and by day 10 was observed in every rat. Harmaline induced tremor appeared later, developing between days 9-12. All rats showed tremor in response to harmaline at day 12 and afterwards. The difference in time of onset between the response to the two drugs reflects different brain sites of action. The development of harmaline induced tremor may reflect the functional synaptic maturation of the olivo-cerebellar circuit.