d -aspartate localizations imply neuronal and neuroendocrine roles

Abstract

Though L-amino acids predominate in living organisms, substantial levels of free D-serine and D-aspartate occur in mammals, especially in nervous and endocrine tissues. Using an antibody specific for glutaraldehyde-fixed D-aspartate, we have localized D-aspartate in rat tissues. In the brain we observe discrete neuronal localizations of Daspartate, especially in the external plexiform layer of the olfactory bulb, hypothalamic supraoptic and paraventricular nuclei, the medial habenula, and certain brainstem nuclei. In rats 3-4 weeks old, we observe D-aspartate in septal nuclei and in a subset of stellate and basket cells of the cerebellum. D-aspartate is also concentrated in glands, including the epinephrine cells of the adrenal medulla, the posterior pituitary, and the pineal gland. Levels in the pineal gland are the highest of any mammalian tissue. Although D-amino acids are well known in bacterial physiology, only recently have they been found in mammals (1-3). Substantial levels of D-serine occur in brain tissue with a regional distribution resembling the the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors (4). By immunohistochemistry we have shown that D-serine is selectively localized to a subpopulation of astrocytes in close proximity to NMDA receptors and released by glutamate stimulation (5, 6). Since D-serine potently activates the glycine site on NMDA receptors, these findings indicate that D-serine is a novel messenger molecule that serves as an endogenous ligand for this site.Lajtha and coworkers (1) discovered very high levels of D-aspartate in the brain and other tissues of mammals. Daspartate levels are highest in neonatal tissues, attaining millimolar concentrations in newborn rat cerebral cortex, pituitary gland, and 3-week-old adrenal gland (7,8). Using immunohistochemistry, we now describe selective neuronal localizations of D-aspartate in discrete brain areas and endocrine structures. D-aspartate oxidase (DAPOX), visualized by a novel histochemical technique, is localized inversely to endogenous D-aspartate. MATERIALS AND METHODSMaterials. Sprague-Dawley rats were purchased from Sasco (Wilmington, MA). Hypophysectomized rats were from Charles River Breeding Laboratories. Mice (129͞SvEv Agouti strain) were from Taconic Farms. Antibody to L-aspartate (9) was from Chemicon. Glutaraldehyde (GA) was from Electron Microscopy Sciences (Fort Washington, PA). The peroxidase Elite staining kit was from Vector Laboratories. All other reagents were from Sigma.Antibody Production and Purification. D-aspartate was coupled to BSA with GA and then reduced with NaBH 4 (10). Rabbits were immunized using a colloidal gold technique (5, 11), and stereoselective, high-affinity antiserum was produced after 7 weeks. Before use, batches of D-aspartate antiserum were diluted 1:10 with 0.05% NaN 3 and 10 mM Tris (pH 7.4). Diluted serum (10 ml) was mixed with 1 ml packed agarose beads that had been coupled to GA-BSA at a concentration of 4.3 mg͞ml with respect to BSA. Antiserum and beads were incubate...

Aspartate racemase, generating neuronal D-aspartate, regulates adult neurogenesis

“…If DR physiologically forms D-aspartate, then the enzyme and its product should have similar localizations. D-aspartate is highly concentrated in the supraoptic and paraventricular hypothalamic nuclei, whose axons terminate in the posterior pituitary (19). Our immunohistochemical studies revealed similarly high densities of DR in the paraventricular and supraoptic nuclei (Fig.…”

“…The free-floating sections were quenched with 0.1% H 2 O 2 in methanol/PBS, blocked with 4% normal goat serum and 0.2% Triton X-100 in TBS (blocking buffer), and incubated with DR antibody, diluted 1:700 in blocking buffer, overnight at 4°C. For immunohistochemistry, DR antibody works optimally D-Aspartate immunohistochemistry was conducted as described previously (19). C57BL/6 mice were perfused with 5% glutaraldehyde/0.5% paraformaldehyde at 37°C.…”

d-Aspartate Prevents Corticostriatal Long-Term Depression and Attenuates Schizophrenia-Like Symptoms Induced by Amphetamine and MK-801

“…Although the function of D-aspartate in mammalian brain is still matter of debate, mounting evidence has indicated that this D-amino acid may act as a putative neuromodulator of the glutamatergic system (Schell et al, 1997;Errico et al, 2008a). Here we examined the hypothesis that elevation of D-aspartate concentrations in adult mice might unveil its central influence in behaviors and electrophysiological paradigms associated with NMDAR signaling.…”

“…Another D-amino acid, named D-aspartate, occurs in the mammalian brain. D-Aspartate levels are high in the embryonic phase and dramatically decrease during postnatal life, in concomitance with increased expression of D-aspartate oxidase (DDO), the enzyme responsible for its deg- radation (Schell et al, 1997;Errico et al, 2006;Huang et al, 2006). In contrast to D-serine, now well characterized for its implication in NMDAR-dependent functions, the role of D-aspartate in the CNS remains elusive.…”

A Role for D-aspartate Oxidase in Schizophrenia and in Schizophrenia-related Symptoms Induced by Phencyclidine in Mice.

“…12 Free D-aspartate (D-Asp) is present in markedly high amounts in the embryonic brain of mammals. After birth, the endogenous levels of this atypical amino acid rapidly decrease, [13][14][15][16] due to the onset of the D-aspartate oxidase (DDO) activity. 17 Recent evidence has shown that D-Asp activates NMDARs through the binding to the glutamate site of GluN2 subunits.…”