Zehava Gottesfeld

Regional Mapping of Neostriatal Neurotransmitter Systems as a Function of Aging

Abstract: The purpose of the present investigation was to map chemically the distribution of certain neurotransmitter systems in the neostriatum of rats aged 6, 16, and 26 months. This mapping was carried out by microdissection of discrete striatal regions coupled with radiometric assays for choline acetyltransferase (ChAT), glutamate decarboxylase (GAD), dopamine (DA), and norepinephrine (NA). In all age groups, ChAT, DA, and NA were highest in the rostral relative to the caudal neostriatum. Additionally, ChAT was higher in the lateral than in the medial region, whereas GAD was more homogeneously distributed within the striatum. ChAT activity was decreased significantly primarily in the caudal regions in rats aged 16 and 26 months. DA levels were decreased in the caudal striatum in rats aged 26 months. NA levels were found to be significantly decreased primarily in the rostral neostriatal regions of the otdest rats. GAD activity remained unchanged in all age groups. These regional changes in selected neurotransmitter systems may underlie specific motor and cognitive deficits that often occur during aging.

Stria medullaris: a possible pathway containing GABAergic afferents to the lateral habenula

The habenular complex (Hb) is an epithalamic structure located bilaterally along the dorsal edge of the third ventricle and consists of medial (MHb) and lateral (LHb) nuclei. The habenula has been associated with the limbic system 24 and functionally linked with a variety of roles related to this system4,5,z0, 30. The forebrain limbic structures are connected to the Hb through one of its major pathways, the stria medullaris (SM). Studies in several mammalian species have demonstrated that fibers from diverse sources pass via the SM and project to the Hb (for review see refs. 6, 17, 23). A paucity of knowledge exists concerning the identity of the neurotransmitter(s) which are released from these fibers. Further information along this line could improve our understanding of the role of the Hb in brain function. The use of histofluorescence, histochemical and sensitive microassay techniques have revealed in the Hb the presence of putative transmitter substances and their associated metabolic enzymes such as catecholamines11, 33, serotoninl, 3, choline acetyltransferase 27, acetylcholinesterase11, 33, glutamic acid decarboxylasO s,32 and substance pg. However, the amount of information on the compartmentalization of these substances in the Hb is relatively small, although the presence of cholinergic and substance P-containing cell bodies in the M H b was recently reported°,12,16, ~1. The present work will focus on the gamma-aminobutyric acid (GABA) system in the Hb, as manifested by the activity of its synthesizing enzyme glutamic acid decarboxylase (GAD), a marker of GABAergic neurons. In the habenular complex, G A D activity is primarily contained in the LHb is and to a large extent, probably, in GABAergic nerve terminals. The latter are characterized in other regions by a high GABA content 25 and G A D activity 7 as well as the capacity to accumulate exogenous GABA by a high affinity sodium and energydependent processes ~0. Afferent projections to the Hb through the SM have been reported to emanate from various regions such as the globus pallidus (GP) and ento-

Maternal and paternal alcohol use : effects on the immune system of the offspring

There is no single mechanism which can account for such a complex biological phenomenon as immune regulation, nor is it clear how alcohol teratogenicity exerts its multiple adversive effects, including lasting immune deficits. Much of the research aimed at unravelling effects of pre- or early postnatal alcohol exposure on the organisms defense mechanisms and long-term health risks has been phenomenological. A better understanding of mechanisms which underlie alcohol effects on immune competency will require integrated studies of the neuro-immune-endocrine networks.

High-affinity uptake of neurotransmitters in rat neostriatum: effects of aging.

High‐affinity uptake of dopamine (DA), glutamate, and γ‐aminobutyric acid (GABA) was determined in crude synaptosomal preparations from neostriatal regions of rats 7, 17, and 27 months of age. Dopamine uptake was highest in rostral neostriatum, but no age‐related differences were detected. On the other hand, the high‐affinity uptake of both GABA and glutamate was increased with age. This may reflect astrocytic hypertrophy or hyperplasia, which have been reported to occur in the neostriatum during the aging process.

Lack of biochemical evidence for a direct habenulo-raphe GABAergic pathway

Morphological observations have suggested that the habenular nuclei, particularly the lateral habenula (LHb), are a major source of afferents to the mesencephalic raphe region. These connections have been demonstrated earlier by a fiber degeneration technique iv, and more recently by autoradiography v and horseradish peroxidase tracing ~,ag,~° (see, however, ref. 16). In a recent electrophysiological study it was reported that habenular stimulation inhibited the firing of cells in the dorsal raphe (dr) nuclei, and that gamma-aminobutyric acid (GABA) might be the neurotransmitter contained within the habenulo-raphe projection el. The present work has been undertaken to biochemically test this suggestion. Glutamic acid decarboxylase (GAD), the GABA synthesizing enzyme, was used as a marker of GABAergic neurons. The activity of GAD in the dr was measured in rats with habenular lesions following different survival periods. Choline acetyltransferase (CHAT), a marker of cholinergic neurons, was measured in the interpeduncular nucleus (ip) of the same animals in order to verify the completeness of the lesion s,ae,14. The results of this study do not support the hypothesis that a direct GABAergic pathway exists between the LHb and the dr nuclei. Male Sprague-Dawley rats weighing about 300 g were obtained from ZivicMiller (Allison Park, Pa.). Bilateral radiofrequency lesions of the habenula were placed in animals anesthetized with Equi-Thesin (Jensen-Salsbery, Kansas City, Mo.). The lesions were made with stainless steel electrodes (0.5 mm in diameter) insulated with Epoxy resin except for 1.5 mm at the tip. The coordinates were according to Kanig and Klippel 1° and adjusted for body weight. The habenular lesion was extensive and included also the emerging part of the fasciculus retroflexus (FR) (Fig. 1) at 4.7

Effects of repeated immobilization stress on glutamate decar☐ylase and choline acetyltransferase in discrete brain regions

A variety of stressors trigger the release of hypothalamic corticotropin releasing factor (CRF) which activates the pituitary-adrenal axis. This action is manifested by the release of adrenocorticotrophic hormone (ACTH) and subsequently of adrenal corticoids. The secretion of CRF is thought to be regulated, at least in part, by a combined facilitatory and inhibitory action, possibly mediated by the amines, acetylcholine and gamma-aminobutyric acid (GABA) (for review see refs. 7 and 21). This stress is also attended by striking CNS-mediated increases in sympathetic-adrenal medullary discharge. The initiation of adrenergic activation by stress involves pathways similar to those controlling hypothalamic releasing factors which control pituitary hormones, and it is likely that similar neurotransmitters are involved. A large number of studies have focused attention on the role of neurotransmitters in mediating the response to stress, but only recently has information been obtained from studies which combine microdissection of discrete brain regions with sensitive microassay techniques. These studies have shown that differential changes of catecholamines and serotonin occur in discrete brain regions in response to various stressors 12, 15,17,1s. Comparable knowledge of the behavior of cholinergic and GABAergic neurons in individual nuclei is not available. Such information is of significance, since it could reveal a functional role of certain discrete brain areas. The purpose of this work was to compare the effect of immobilization stress on glutamate decarboxylase (GAD) and choline acetyltransferase (CHAT), marker enzymes of GABAergic and cholinergic neurons, respectively, in discrete brain regions, and to study the sequence of changes that occur after short-term as well as prolonged stress when adaptation is likely to occur. The results demonstrate differential changes in GAD and ChAT activity in specific nuclei. Male Sprague-Dawley rats (300-350 g, specific pathogen-free) purchased from Taconic Farms were used. Food and water was provided ad libitum. The animals were

Distribution of glutamic acid decar☐ylase in certain rhombencephalic and thalamic nuclei of the rat

L-Glutamic acid decarboxylase (GAD) catalyzes the biosynthesis of gammaaminobutyric acid (GABA) from L-glutamic acid. Although GABA is a potential inhibitory neurotransmitter in the mammalian CNSS,9,15,16, relatively little is known about its localization in discrete neuronal pathways in the brain. The best documented of such GABAergic paths are those from cerebellar Purkinje cells to the deep cerebellar and lateral vestibular nucleP s, and the strio-nigral 4 or pallido-nigraP 1 paths. The purpose of this study was to map the distribution of GAD in individual nuclei of the rat brain in order to pursue our longterm goal of identifying GABAergic neuronal pathways in the CNS. Male Sprague-Dawley rats weighing about 250 g and maintained on standard rat chow and water ad lib were killed by decapitation and their brains rapidly removed and frozen on dry ice. Individual nuclei were removed from frozen 300/zm cryostat sections using the microdissection technique of Palkovits la. Fig. 1 shows the sites where tissue samples were removed. Bilateral tissue samples were placed in 25 #l of ice-cold 10 m M EDTA pH 7.3, containing 0.2 ~ Triton X-100, and homogenized by sonication 6. A 5 #l aliquot was immediately taken for protein analysis according to the micromethod of Lowry et al.10. The remaining samples were stored frozen until assayed. GAD activity was determined by a sensitive radiometric method 1, as modified by Fonnum et al. a, except that the final glutamate concentration in the incubation medium was 10 mM. The reaction was initiated by adding 2 #l of the substrate mixture a to 2 #l of tissue homogenate, containing 0.2-2.0/~g protein. Duplicate samples were incubated at 37 °C for 60 min. The reaction was stopped by injecting excess 0.5 N H2SO4 into the incubation medium. The samples were then further incubated for 45 min, and the 14COz released was trapped in 50 #1 of hyamine. The hyamine-tubes were then placed in a counting vial containing 1 ml of absolute ethanol and l0 ml of scintillator (6.4 g PPO and 75 rng POPOP per liter toluene) and counted in a liquid scintillation spectrometer. Blanks

Regional Distribution of Catecholamines in Nucleus Accumbens of the Rabbit

Abstract: The nucleus accumbens is an important telencephalic region, which is the target limbic and mesolimbic pathways. Because of an ongoing physiological study of the effects of dopamine, we wanted to determine regional differences of dopamine and norepinephrine concentrations in the nucleus. As determined by radioenzymatic assays, dopamine levels were not significantly different in the anterior‐posterior dimension, averaging approximately 187 ng dopamine/mg protein. Substantial amounts of norepinephrine were found throughout the nucleus, but the levels were significantly higher in the caudal portions of the nucleus, being approximately 4.5 times higher than in the anterior portions.

Origin and distribution of noradrenergic innervation in the habenula: A neurochemical study

The distribution of norepinephrine (NE) was chemically mapped in discrete regions of the rat habenula (Hb). In addition, the contribution of both peripheral sympathetic and central noradrenergic sources to habenula NE was quantified following selective deafferentations. It was found that NE is distributed hetereogeneously in discrete Hb regions. Furthermore, it was demonstrated that after superior cervical ganglionectomy a small (26%) but significant (P less than 0.05) decrease in NE occurred only in the medial Hb (MHb), while NE in the lateral Hb (LHb) remained unaltered. In contrast, deafferentation of central noradrenergic innervation, such as lesions in the ascending dorsal noradrenergic bundle (DNB), caused a marked decrease in NE in both MHb (53%) and LHb (50%). Lesioning the dorsal longitudinal fasciculus resulted in NE loss in both MHb and LHb by 31%. These data corroborate the histofluorescent demonstration of the noradrenergic innervation of the Hb.

Developmental delays associated with prenatal alcohol exposure are reversed by thyroid hormone treatment

Exposure to alcohol in utero has been associated with hypothyroidism and a variety of developmental defects characteristic of thyroid dysfunction. The present work examined whether these abnormalities could be reversed in infant rats treated with thyroid hormones. Subjects were offspring of dams which were on the following diet regimen during gestation: (1) free access to liquid diet containing ethanol (alcohol pups); (2) an equal volume of isocaloric liquid diet (pair-fed pups); or (3) ad libitum control diet (control pups). Neonates from each group were foster-nursed by control dams, and received triiodothyronine (T3; 0.1 mg/kg/day; s.c.) or saline treatments on postnatal days 1 to 10. The alcohol neonates displayed reduced serum thyroxine which was restored to normal by postnatal day 14. In addition, these pups showed a delayed appearance of developmental landmarks, including righting reflex, dental eruption, auditory startle response and eye opening. The retarded incisor eruption and eye opening were reversed in alcohol pups by T3 treatments. The present data suggest that at least some of the developmental abnormalities associated with prenatal alcohol exposure are attributable to perinatal hypothyroidism and can be restored by early hormone replacement therapy.