Genetic deletion of angiotensin AT2 receptor leads to increased cell numbers in different brain structures of mice

doi:10.1016/S0167-0115(01)00258-0

Regulatory Peptides

Volume 99, Issues 2–3, 15 June 2001, Pages 209-216

https://doi.org/10.1016/S0167-0115(01)00258-0 Get rights and content

Abstract

Angiotensin II (Ang II) is a potent vasoactive peptide and displays growth factor-like properties. Different high-affinity Ang II receptor subtypes (AT1_A, AT1_B and AT2) have been cloned. They are expressed in various brain structures. Additionally, it has been assumed that Mas could interact directly or indirectly with the renin–angiotensin system. The AT1 receptor mediates pressor and mitogenic effects of Ang II, whereas physiological function and signaling mechanisms of the AT2 receptor remain poorly understood. Recent reports have shown that Ang II could mediate apoptosis through AT2 receptors. Since the AT1_A, AT2 and Mas knockout mice provide new tools for uncovering potential actions of Ang II, the cell number in different brain structures of male adult wild-type mice and mice deficient for AT1_A, AT2 or Mas was evaluated to get more insight into the role of Ang II in central nervous system development. In nearly all investigated brain structures (cortex, hippocampus, amygdala, thalamus), the cell number was significantly higher in AT2-deficient mice in comparison to wild-type mice. To the contrary, in AT1_A-deficient mice the cell number was significantly less than in controls in the lateral geniculate and the medial amygdaloid nucleus. However, cell numbers were not changed in Mas-knockout mice compared to their wild-types. These results show the contrary effects of both angiotensin receptors on cell growth and represent the first demonstration of their action on neuronal cell development evidenced in the adult mouse brain.

Introduction

The different actions of Ang II are partly due to the diversity of its receptors (AT1 and AT2). The AT1 receptor was shown to consist of two isoforms (AT1_A and AT1_B) in rodents, whereas only one AT1 receptor was found in higher mammals. Most of the known Ang II actions on cardiovascular regulation, hormone secretion and fluid balance are mediated by AT1 receptors. In contrast, the physiological actions of the AT2 receptor remain largely uncertain. The AT2 receptor is highly expressed in neonatal tissue. AT2 levels decrease sharply after birth, but persist in adulthood in the brain [1], which leads to the assumption that the AT2 receptor might be involved in differentiation, development or apoptosis [2].

Recent in vitro studies have shown that Ang II induces apoptosis in different cell types [3], [4], [5], [6] through AT2 receptors. Additionally, it has been recently demonstrated that Ang II, via AT2 receptors and activation of a serine/threonine phosphatase, contributed to apoptosis in neurons cultured from newborn rat hypothalamus and brain stem [7]. Moreover, it could be shown that the AT2 receptor mediates vascular smooth muscle cell apoptosis, whereas the AT1 receptor stimulation in vitro leads to a counteracting effect [8].

The Mas protooncogene is a gene, encoding an orphan G-protein coupled receptor. Mas is expressed predominantly in testis and forebrain and its expression in the brain is developmentally regulated [9]. Ang II has been suggested to be a ligand for Mas because of the Ang II-induced accumulation of inositol phosphates and calcium in Mas-transfected cells [10]. It has been shown that Mas enhances the effects of Ang II on cells expressing endogenously the AT1 receptor [11].

Mutant mice with targeting deletion of one of both angiotensin receptors or of the Mas protooncogene have been recently produced. The AT1_A knockout mice (AT1-KO) [12] develop and grow up normally [13], but exhibit chronic hypotension and renin overproduction [14]. AT2 knockout mice (AT2-KO) also develop normally, but exhibit decreased spontaneous movements and exploratory behavior as well as impaired drinking responses to water deprivation compared to wild-type mice [15], [16]. Blood pressure and sodium excretion showed a sustained hypersensitivity to Ang II [17]. Mas-knockout mice have been recently produced, and these mice develop and breed normally. However, hippocampal long-term potentiation (LTP) and anxiety are markedly affected [18]. Our recent results have supported findings concerning an interaction of Mas and the AT1 receptor [19].

The aim of the present study was the morphometric evaluation of the cell number in different brain structures of wild-type mice and mice with a deletion of Ang II interacting receptors (AT1-, AT2-and Mas-KO) to get a first insight into the possible functions of Ang II in the development of the brain.

Section snippets

Materials and methods

All procedures were carried out according to the standards of animal welfare and approved by the regional Berlin Animal Ethics Committee (TO 320/97).

Adult male mice in an age of 14 weeks, with a deletion of the Mas protooncogene (Mas^−/−) [18], angiotensin receptor AT1_A gene (AT1_A^−/−) [12], or angiotensin receptor AT2 gene (AT2^−/y) [16] and their own control strains (Mas^+/+; AT1_A^+/+; AT2^+/y) were used for the experiments. The animals were maintained under standardized conditions with an

Nomenclature

The anatomical localization for each brain nucleus was verified by the help of a stereotaxic atlas of the mouse brain [21]. The nomenclature is based on the nomenclature and abbreviations used in this atlas (see also Fig. 2).

Effects of genetic deletions on neuronal cell number

The somatosensory and the piriform cortex were chosen to analyze possible changes in cortical areas in the three knockout models. Fig. 3 illustrates the distribution of cells in different cortex areas of wild-type (control), AT2-, AT1- and Mas-KO. In contrast to the

Discussion

A recent study using angiotensinogen-knockout mice revealed a decreased density in cells of hippocampus and cerebellum, suggesting that the renin–angiotensin system could play a critical role in central nervous system development, preventing neuronal cells from apoptosis [22].

Our study provides new information about the ability of angiotensin receptors to control processes occurring presumably during development, which led to changes in the adult brain. Candidate mechanisms might be alterations

Acknowledgments

We thank Dr. T.M. Coffman for providing the AT1_A-knockout mice and Dr. Volkmar Groß for providing breeding pairs of AT2 knockout mice. We also thank Dr. M.J. Wayner, Dr. D. Armstrong and Dr. C. Phelix for critically reading of the manuscript and helpful discussion.

This study was supported by the Deutsche Forschungsgemeinschaft (Al 342/7-1).

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Genetic deletion of angiotensin AT2 receptor leads to increased cell numbers in different brain structures of mice

Abstract

Introduction

Section snippets

Materials and methods

Nomenclature

Effects of genetic deletions on neuronal cell number

Discussion

Acknowledgments

J. Biol. Chem.

Brain Res. Dev. Brain Res.

Biochim. Biophys. Acta

Kidney Int.

J. Biol. Chem.

Biochim. Biophys. Acta

Neurosci. Lett.

Biochem. Biophys. Res. Commun.

Brain Res.

Brain Res. Dev. Brain Res.

Mol. Cell Endocrinol.

Neuropeptides

Brain Res.

Ontogeny of angiotensin II type 2 receptor mRNA expression in fetal and neonatal rat brain

J. Comp. Neurol.

Characterization and development of angiotensin II receptor subtypes (AT1 and AT2) in rat brain

Am. J. Physiol.

Angiotensin II type 2 receptor mediates programmed cell death

Proc. Natl. Acad. Sci. U. S. A.

Angiotensin II type 2 receptors mediate inhibition of mitogen-activated protein kinase cascade and functional activation of SHP-1 tyrosine phosphatase

Biochem. J.

Angiotensin II induces apoptosis of human endothelial cells

Circ. Res.

Angiotensin II type 2 receptor-mediated apoptosis of cultured neurons from newborn rat brain

Endocrinology

Angiotensin II type 2 receptor mediates vascular smooth muscle cell apoptosis and antagonizes angiotensin type 1 receptor action: an in vitro gene transfer study

Life Sci.

The mas oncogene encodes an angiotensin receptor

Nature