Deficits in nitric oxide production after tetanic stimulation are related to the reduction of long‐term potentiation in Schaffer‐CA1 synapses in aged Fischer 344 rats

In the present study, we investigated whether nitric oxide (NO) production after tetanic stimulation is involved in long‐term potentiation (LTP) in Schaffer‐CA1 synapses in both young adult and aged rats. The changes in both the population spike amplitude and NO metabolites, nitrite (NO₂^–) and nitrate (NO₃^–), in the CA1 region were simultaneously determined before and after tetanic stimulation. Increases in NO_x (NO₂^– plus NO₃^–) levels in the CA1 region were observed after tetanic stimulation in young adult rats as well as increase in the population spike amplitude. In aged rats, LTP was significantly inhibited compared with that in young adult rats. No increase in NO_x level after tetanic stimulation was observed in aged rats. These findings directly demonstrated that NO production might be involved in the process of LTP formation in Schaffer‐CA1 synapses of the rat hippocampus, and that the deficiency of hippocampal NO production might be responsible for reduction of LTP formation in aged rats.     

Deficits across multiple cognitive domains in a subset of aged Fischer 344 rats

Rodent models of cognitive aging routinely use spatial performance on the water maze to characterize medial temporal lobe integrity. Water maze performance is dependent upon this system and, as in the aged human population, individual differences in learning abilities are reliably observed among spatially characterized aged rats. However, unlike human aging in which cognitive deficits rarely occur in isolation, few non-spatial learning deficits have been identified in association with spatial impairment among aged rats. In this study, a subset of male aged Fischer 344 rats was impaired both in water maze and odor discrimination tasks, whereas other aged cohorts performed on par with young adult rats in both settings. The odor discrimination learning deficits were reliable across multiple problems. Moreover, these deficits were not a consequence of anosmia and were specific to olfactory learning, as cognitively impaired aged rats performed normally on an analogous non-olfactory discrimination task. These are among the first data to describe an aging model in which individual variability among aged rat cognition occurs across two independent behavioral domains.     

Decreased sympathetic innervation of spleen in aged Fischer 344 rats

Splenic noradrenergic innervation in young adult and aged Fischer 344 rats was examined using fluorescence histochemistry for catecholamines and high performance liquid chromatography with electrochemical detection (LCEC) for the quantitation of norepinephrine (NE). In young adult rats, abundant noradrenergic plexuses followed the vasculature and trabeculae into splenic white pulp. In aged rats, noradrenergic innervation was reduced in density and in overall intensity of fluorescence, and splenic NE levels were significantly lower. The relationship between diminished noradrenergic innervation and diminished immune responsiveness in aging mammals, while not clear on a causal level, is presented as a hypothesis for further testing.     

Blockade of hippocampal long-term potentiation by sustained tetanic stimulation near the recording site

Specific patterns of electrical stimulation trigger several forms of synaptic plasticity in hippocampal pyramidal cells, including a long-term potentiation (LTP) of excitatory synaptic transmission. I investigated the effect of commonly used stimulation protocols at different distances from the recording site. Sustained electrical stimulation (100 Hz, 1 s) delivered close to the recording site prevented LTP induction; the same stimulation from a second electrode placed farther away subsequently produced LTP at the same recording site. Strong stimulation near the recording site could also interfere with LTP triggered from a distal site. In contrast to sustained high-frequency stimulation, intermittent stimulation (theta burst pattern) delivered close to the recording site produced normal LTP. These data support the hypothesis that strong stimulation releases a factor that acts locally to prevent LTP.     

Stress-induced changes in brain-derived neurotrophic factor expression are attenuated in aged Fischer 344/N rats

Aging and stress can sometimes result in a decline in brain function. We addressed the question whether changes in the expression of neurotrophic factors, which are necessary for the survival and maintenance of neurons, might occur during aging and stress. Therefore, we used in situ hybridization to investigate the effects of aging and stress on neurotrophic factor expression in young (3–4 month) and old (24 month) male Fischer 344/N rats. The ability of acute immobilization stress (2 h) to modulate BDNF mRNA levels in old rats was significantly reduced both in the hippocampus (a smaller decrease in BDNF) and the PVN (a smaller increase in BDNF) compared to young rats. In contrast, the induction of nerve growth factor and neurotrophin 3 (NT-3) by stress was not influenced by age. The diminished BDNF responses to stress in aged rats may be relevant to difficulties in adaptation to stress encountered during old age.     

Hippocampal NGF levels are not reduced in the aged Fischer 344 rat.

The sympathetic sprouting response that occurs in the rat hippocampal formation following septal denervation is reduced in aged rats. Since considerable evidence implicates NGF-like activity in eliciting the sprouting, the simplest explanation for the age-related decline in sympathetic sprouting is a reduction in hippocampal NGF levels. In the present study, hippocampal NGF levels were measured using a 2-site ELISA in four different age groups of Fischer 344 rats. There was no decline in NGF levels with age, nor did we find any differences between male and female rats. This contradicts an earlier report in which a 40% reduction in hippocampal NGF protein levels was found in aged rats. Possible reasons for this discrepancy are discussed. The present results do not support the hypothesis that the age-related decline in sympathetic sprouting is due to a reduction in total hippocampal NGF levels.     

Enhancement of long-term potentiation at CA1-subiculum synapses in MK-801-treated rats

The subiculum plays a key role in processing neuronal information from the hippocampus to different cortical and subcortical brain regions. The subicular projections to the nucleus accumbens and the prefrontal cortex have received increasing attention, as alterations of their activity seem to be involved in schizophrenia. Phencyclidine and other non-competitive antagonists of NMDA receptors (such as ketamine and MK-801) induce psychotic effects in humans that closely resemble the positive, negative and cognitive symptoms of schizophrenia. Using the MK-801 model of psychosis, we investigated the time course of alterations of synaptic transmission and plasticity at CA1-subiculum synapses of hippocampal brain slices 4 h, 24 h and 4 weeks after MK-801 treatment. We report here that systemic application of MK-801 causes a facilitation of LTP at CA1-subiculum synapses 24 h after treatment as compared with control LTP. Four weeks after MK-801 treatment, the magnitude of LTP reversed to control values. The priming of LTP 24 h after systemic application of MK-801 suggest a new form of metaplasticity that sheds light on the delayed facilitating effect of this drug on synaptic efficacy.     

Cardiovascular responses to acute footshock stress in adult and aged Fischer 344 male rats

Tail artery catheters were surgically implanted in Fischer 344 male rats to allow for measurement of mean arterial pressure (MAP, mm Hg) and heart rate (HR, beats/min) in conscious, unrestrained rats. Basal values of MAP and HR were similar for groups of 4, 12 and 24 month old rats. Increments in Map did not differ among rats of the 3 ages following handling and transfer to a shock chamber or immediately or 5 minutes after exposure to inescapable footshock (2.0 mA, 0.6 sec duration, every 6 sec for 1 min). In contrast, there was a significant age-related attenuation of the tachycardia following handling and transfer of rats to the shock chamber and at the end of footshock. These data are consistent with previous findings of a reduced sensitivity of the aged myocardium to stress-induced sympathetic stimulation.     

Light intensity-associated eye lesions of Fischer 344 rats in long-term studies.

Albino rats and mice are sensitive to light and the recommended illumination of animal rooms (75-125 ft-candles) is known to cause retinal damage. When a room is illuminated by ceiling lights, animals in the cages of the top row and, to some extent, in the side columns of cage racks will be exposed to higher light intensity than those in the other cages of the rack. In 2-yr chemical carcinogenicity studies of the National Toxicology Program (previously the Carcinogenicity Bioassay Program of the National Cancer Institute), Fischer 344 rats were group-housed in hanging drawer-type clear polycarbonate cages. During the course of the chronic studies, a number of rats developed opacity of the eye. Ocular examination indicated chronic uveitis, deep interstitial keratitis, cataract formation leading to panophthalmitis, and in severe cases, phthisis bulbi. Histologic examination showed cataract and retinal degeneration. Incidences of these lesions were highest (greater than 55%) in the rats of the top rows and lowest in those of the bottom rows (less than 10%) of cages with no relation to chemical treatment, indicating an association with light intensity. The incidence of these eye lesions was markedly decreased (less than 15%) by decreasing the light intensity of the animal room to less than 50 ft-candles at 5 ft from the floor and rotating the cages in each column of a rack from top to bottom when cages or racks were changed.     

Effects of dietary taurine supplementation or deprivation in aged male Fischer 344 rats

Taurine is a sulfur amino acid that is present in high concentration in mammalian tissues and previously has been reported to decline in a number of tissues with advancing age. The aims of the present study were to examine: (1) the effects of dietary taurine supplementation; (2) the effects of taurine-free diets; (3) the ability of aged rats to conserve urinary taurine; and (4) the consequences of these dietary manipulations on some biochemical parameters. Male F344 rats (n = 30/group) 18 months of age were placed on control diets, diets supplemented with 1.5% taurine in the drinking water, or a taurine-free diet for 10 months. An adult control group (12 months old at the end of the study) on normal diets was included for comparison purposes. Significant (P < 0.05) age-related declines in taurine content were observed in the spleen, kidney, eye, cerebellum and serum. Taurine supplementation corrected these deficits in tissue content in aged rats and in many cases increased taurine content above that of adult controls. Urinary excretion of taurine was significantly (P < 0.05) reduced in aged rats indicating an increased need to conserve taurine. Taurine-deficient diets did not further exacerbate the age-related decline in tissue taurine content, suggesting biosynthetic adaptations to the lack of dietary taurine. Dietary taurine supplementation blunted age-related declines in serum IGF-1 and increases in serum creatinine and blood urinary nitrogen (BUN). These studies suggest that advanced aging results in a taurine-deficient state that can be corrected by dietary supplementation.     

Macrophages are unsuccessful in clearing aggregated alpha‐synuclein from the gastrointestinal tract of healthy aged Fischer 344 rats

With age, alpha‐synuclein (α‐SYNC) misfolds and forms insoluble deposits of protein in the myenteric plexus, leading presumably to dystrophy and degeneration in the circuitry controlling gastrointestinal (GI) function. The present experiment examined aggregates of α‐SYNC in the aging small intestine and investigated how macrophages in the wall of the GI tract respond to these aberrant deposits. Groups of adult and aged Fisher 344 rats were studied. Whole mounts of duodenal, jejunal, and ileal smooth muscle wall, including the myenteric plexus, were prepared. Double labeling immunohistochemistry was used to stain α‐SYNC protein and the phenotypic macrophage antigens CD163 and MHCII. Alpha‐synuclein accumulated in dense aggregates in axons of both postganglionic and preganglionic neurons throughout the small intestine. Staining patterns suggested that deposits of protein occur initially in axonal terminals and then spread retrogradely toward the somata. Macrophages that were adjacent to dystrophic terminal processes were swollen and contained vacuoles filled with insoluble α‐SYNC, and these macrophages commonly had the phenotype of alternatively activated phagocytes. The present results suggest that macrophages play an active phagocytotic role in removing α‐SYNC aggregates that accumulate with age in the neural circuitry of the gut. Our observations further indicate that this housekeeping response does not clear the protein sufficiently to eliminate all synucleinopathies or their precursor aggregates from the healthy aging GI tract. Thus, accumulating deposits of insoluble α‐SYNC in the wall of the GI tract may contribute, especially when compounded by disease or inflammation, to the age‐associated neuropathies in the gut that compromise GI function. Anat Rec, 296:654–669, 2013. © 2013 Wiley Periodicals, Inc.     

Quantitation of synaptic density in the septal nuclei of young and aged Fischer 344 rats

Synaptic density in the medial and lateral septal nuclei was examined in 3 and 24-28 months of age Fischer 344 rats. The lateral nucleus had a higher synaptic density than the medial region in both age groups. There were no statistically significant differences in synapse density in either region as a function of age, but the data suggested a subpopulation of aged animals which did show an age-related decline in synaptic density in the lateral, but not medial area of the septum. These data indicate that sample size may be an important variable in assessing possible age-related differences in synaptic density, since a broad range of values, some significantly below the range of young animals, exists in the aged brain.     

Role of nitric oxide in long-term potentiation of the rat medial vestibular nuclei

In rat brainstem slices, we investigated the role of nitric oxide in long-term potentiation induced in the ventral portion of the medial vestibular nuclei by high-frequency stimulation of the primary vestibular afferents. The nitric oxide scavenger [2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide ] and the nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester were administered before and after induction of potentiation. Both drugs completely prevented long-term potentiation, whereas they did not impede the potentiation build-up, or affect the already established potentiation. These results demonstrate that the induction, but not the maintenance of vestibular long-term potentiation, depends on the synthesis and release into the extracellular medium of nitric oxide. In addition, we analysed the effect of the nitric oxide donor sodium nitroprusside on vestibular responses. Sodium nitroprusside induced long-term potentiation, as evidenced through the field potential enhancement and unit peak latency decrease. This potentiation was impeded by D, L-2-amino-5-phosphonopentanoic acid, and was reduced under blockade of synaptosomal platelet-activating factor receptors by ginkgolide B and group I metabotropic glutamate receptors by (R,S)-1-aminoindan-1, 5-dicarboxylic acid. When reduced, potentiation fully developed following the washout of antagonist, demonstrating an involvement of platelet-activating factor and group I metabotropic glutamate receptors in its full development. Potentiation induced by sodium nitroprusside was also associated with a decrease in the paired-pulse facilitation ratio, which persisted under ginkgolide B, indicating that nitric oxide increases glutamate release independently of platelet-activating factor-mediated presynaptic events.We suggest that nitric oxide, released after the activation of N-methyl-D-aspartate receptors, acts as a retrograde messenger leading to an enhancement of glutamate release to a sufficient level for triggering potentiation. Once the synaptic efficacy has changed, it becomes a long-lasting phenomenon only through a subsequent action of platelet-activating factor.     

Deficits in the expression of the NR2B subunit in the hippocampus of aged Fisher 344 rats

The NMDA receptor (NMDAR) has been implicated in the induction of LTP at hippocampal synapses, and has been proposed to play a significant role in the involvement of the hippocampus with learning and memory. Aged rats are known to have deficits in LTP, learning and memory. We tested the hypothesis that aged rats might have deficits in expression of NMDAR subunits. Aged rats have significantly lower levels of NR2B mRNA and protein compared to young animals. This complements a recent report which showed improved learning and memory in mice which overexpress NR2B. No changes were seen in either the mRNA or the protein levels of the NMDAR subunit NR2A, nor in the alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionate receptor (AMPAR) subunit GluR2. Our data support the hypothesis that age related alterations in the expression of the NMDAR NR2B subunit might underlie deficits in LTP and learning and memory in aged animals.     

Cellular kinetics in the lungs of aging Fischer 344 rats after acute exposure to ozone.

Lung repair in aging Fischer 344 male rats was investigated after an acute inhalation exposure to ozone. Adult (9-month-old) and senescent (24-month-old) rats were exposed to 0.8 ppm ozone for a single period of 6 hours, and thereafter studied over 5 days of recovery in clean air. The animals were given intraperitoneal injections of colchicine and [3H]thymidine, 4 hours and 1.5 hours before termination, respectively. The lungs were inflated with glutaraldehyde, and tissue samples were embedded in epoxy resin for electron microscopy, or in glycol methacrylate for light-microscopic autoradiography. Exposure to ozone produced epithelial injury in alveolar ducts and terminal bronchioles, later reflected by the transient increase in mitotic activity of nonciliated bronchiolar cells and alveolar type 2 cells. The increase in metaphase-arrested cells and [3H]thymidine-labeled cells in bronchioles followed similar time courses, ie, maximal at days 1.5 to 2, and subsiding by day 3. In the alveoli, type 1 cell necrosis was observed early after exposure (6 hours recovery), without notable structural changes in the interstitial and endothelial compartments. The increased mitotic activity in the alveolar septa was mostly due to proliferation of epithelial type 2 cells, which was maximal at day 1.5, and of interstitial cells, maximal at day 2.5. The magnitude of the mitotic responses of nonciliated bronchiolar cells, alveolar type 2 cells and interstitial cells was highest (+50%) in the lungs of senescent rats. Although the cellular events during repair are essentially similar in both age groups, the results indicate that senescent rats have a significantly higher level of initial injury from inhalation of ozone than adult animals.     

Impairment of neocortical long-term potentiation in mice deficient of endothelial nitric oxide synthase

The role of the possible retrograde messenger nitric oxide (NO) in the induction of long-term potentiation (LTP) was studied in supragranular layers of somatosensory cortical slices obtained from adult mice. High-frequency stimulation produced a slowly rising, long-lasting (50 min) and significant (P < 0.001) increase in the extracellular synaptic response by 23%. The induction of LTP was independent from activation of N-methyl-D-aspartate (NMDA) receptors, but prevented by bath application of NG-nitro-L-arginine methyl ester (L-NAME), indicating that one or several of the different NO synthases (NOS) produced NO within the postsynaptic neuron. No LTP could be induced in knockout mice lacking the endothelial NOS (eNOS) isoform. These data suggest that eNOS is involved in an NMDA receptor-independent form of LTP in the rodent cerebral cortex.     

Enhanced hippocampal long-term potentiation in rats after chronic exposure to homocysteine

Homocysteine (HCY) is a sulphur-containing amino acid, which has been linked to neurodegenerative diseases such as Alzheimer's disease, and is widely reported to enhance vulnerability of neurons to oxidative, excitotoxic and apoptotic injury via perturbed calcium homeostasis, activation of N-methyl-D-aspartate (NMDA) and metabotropic glutamate (mGlu) receptors. The present study was undertaken to investigate the effects of HCY on long-term potentiation (LTP) and synaptic transmission after chronic 4-week systemic exposure to HCY in adult rats, and possible longer-term effects of HCY 4 weeks after exposure had ended. Contrary to expectation, LTP was enhanced, not retarded after chronic HCY exposure relative to controls. Basic synaptic transmission was not affected at this time point. However, after the 4-week wash out period, a decrease in speed of basic synaptic transmission emerged, and LTP was still partially enhanced, particularly for time points >30 min post-tetanus. In summary, we provide first evidence for sustained HCY-induced changes in hippocampal plasticity and a slow-onset disruption in synaptic transmission. These changes may reflect the suggested (excito-)toxicity of HCY and its putative contribution to neurodegenerative disease.     

Good things come to those who wait: attenuated discounting of delayed rewards in aged Fischer 344 rats

The ability to make advantageous choices among outcomes that differ in magnitude, probability, and delay until their arrival is critical for optimal survival and well-being across the lifespan. Aged individuals are often characterized as less impulsive in their choices than their young adult counterparts, demonstrating an increased ability to forgo immediate in favor of delayed (and often more beneficial) rewards. Such "wisdom" is usually characterized as a consequence of learning and life experience. However, aging is also associated with prefrontal cortical dysfunction and concomitant impairments in advantageous choice behavior. Animal models afford the opportunity to isolate the effects of biological aging on decision-making from experiential factors. To model one critical component of decision-making, young adult and aged Fischer 344 rats were trained on a two-choice delay discounting task in which one choice provided immediate delivery of a small reward and the other provided a large reward delivered after a variable delay period. Whereas young adult rats showed a characteristic pattern of choice behavior (choosing the large reward at short delays and shifting preference to the small reward as delays increased), aged rats maintained a preference for the large reward at all delays (i.e., attenuated "discounting" of delayed rewards). This increased preference for the large reward in aged rats was not due to perceptual, motor, or motivational factors. The data strongly suggest that, independent of life experience, there are underlying neurobiological factors that contribute to age-related changes in decision-making, and particularly the ability to delay gratification.     

Vasopressin in aged rats: longitudinal studies of vasopressin excretion in Sprague-Dawley and Fischer 344 strains

In order to provide physiological baseline values for future experimental procedures, indices of vasopressin secretion were assessed in male Sprague-Dawley (SD) and Fischer 344 (F344) rats at 3 and 20 months of age. Daily water intake, urine volume, urine osmolality, and urinary vasopressin excretion were monitored in SD rats for 30 days, and in F344 rats for 60 days. In the SD strain, daily water and urine volumes, expressed as ml/24 hr/100 g b.wt., were consistently lower in aged animals, as was a calculation of water balance (water intake-urine output volumes/24 hr). Although mean VP concentration in urine appeared higher in aged rats (33.9 +/- 20.4 pg/ml) than in young (16.3 +/- 7.7 pg/ml), total daily VP excretion was comparable for both ages when expressed as a function of body weight [80.6 +/- 37.3 pg for 3 months old (m.o.) and 81.9 +/- 47.2 pg/24 hr/100 g b.wt. for 3 and 20 m.o. respectively]. Young and old F344 males showed comparable daily drinking and urine volumes, and water balance, during two months of monitoring, but VP excretion was lower (p less than 0.025) in aged rats (83.8 +/- 19.0 pg/24 hr/100 g b.wt.) than in 3 m.o. rats (213.0 +/- 48.1 pg/24 hr/100 g b.wt.). Urine VP concentration was comparable (69.6 +/- 20.6 for 3 m.o.; 59.8 +/- 25.6 pg/ml for 20 m.o.). Mean urine osmolality was not significantly different among groups. Urine osmolality and daily urine volumes showed a significant correlation with daily VP excretion among young, but not aged, rats of both strains.(ABSTRACT TRUNCATED AT 250 WORDS)