Giovanni Diana

Enhancement of learning and memory after activation of cerebral Rho GTPases.

The mechanism whereby the morphology and connectivity of the dendritic tree is regulated depends on an actin dynamics that, in turn, is controlled by Rho GTPases, a family of small GTP-binding proteins encompassing Rho, Rac, and Cdc42 subfamilies. Cytotoxic necrotizing factor 1 (CNF1), a protein toxin from Escherichia coli, constitutively activates Rho GTPases, thus leading to remodeling of the actin cytoskeleton in intact cells. Here, we show that the modulation of cerebral RhoA and Rac1 activity induced by CNF1 in mice leads to (i) rearrangement of cerebral actin cytoskeleton, (ii) enhanced neurotransmission and synaptic plasticity, and (iii) improved learning and memory in various behavioral tasks. The effects persist for weeks and are not observed in mice treated with a recombinant CNF1, in which the enzymatic activity was abolished by substituting serine to cysteine at position 866. The results suggest that learning ability can be improved through pharmacological manipulation of neural connectivity.

Age and strain differences in rat place learning and hippocampal dentate gyrus frequency-potentiation.

Induction of post-tetanic potentiation (PTP) and long-term potentiation (LTP) was analyzed in hippocampal slices obtained from (i) young (6 months old) rats of different strains (Sprague-Dawley, SD; spontaneously hypertensive rats, SHR; and Wistar-Kyoto, WKY), and (ii) from aged (20-24 months old) SD and Fischer 344 (F 344) rats, each group showing a different performance in the Morris maze test. After the application of an electrical tetanus (1 s, 100 Hz, 50 microA) in the stratum moleculare, a significant difference was found in the percent of induction of the dentate PTP in hippocampal slices obtained from rats of different strains and ages. In particular, the induction of the dentate PTP was significantly (P < 0.01) higher in slices obtained from young SD or spontaneously SHR rats, having the better performance in the Morris maze than in slices obtained from old SD or F 344 rats or young WKY rats which had poorer performances in the Morris maze. On the contrary, no significant differences were found in the percent of induction of the LTP in the dentate area of hippocampal slices obtained from rats of different strains and ages. Moreover, after the application of an electrical tetanus (1 s, 100 Hz, 50 microA) in the stratum radiatum, no significant differences were found in the percent of induction of both PTP and LTP in the CA1 area of hippocampal slices obtained from rats of different strains and ages.(ABSTRACT TRUNCATED AT 250 WORDS)

Local cerebral glucose utilization following unilateral and bilateral lesions of the nucleus basalis magnocellularis in the rat

To investigate to what extent the loss of cholinergic projections to the neocortex results in functional impairment in the target areas, local rates of cerebral glucose utilization were measured following excitotoxin lesions of the nucleus basalis magnocellularis (NBM) in the rat. Both unilateral and bilateral lesions of NBM resulted in reversible depression of cerebral metabolism. The effects of unilateral lesions were limited to the cortical areas which receive most of the cholinergic projections from NBM. The metabolic defect produced by bilateral lesions was spread to the whole brain. Within 4 months, however, normal metabolic values coexisted with marked changes of the presynaptic cholinergic markers and impairment of conditioned behavior.

Reduced hippocampal CA1 Ca2+-induced long-term potentiation is associated with age-dependent impairment of spatial learning

Expression of Ca(2+)-induced CA1 long-term potentiation (LTP) was analysed in hippocampal slices obtained from (1) 3-month-old and (2) 18-20-month-old Sprague-Dawley rats selected for their performances in the Morris water maze task. In all slices, a transient (10 min) increase of extracellular Ca2+ concentration (4 mM) caused a long-lasting enhancement of potentials evoked by electrical stimulation of radiatum fibers. However, a significant difference was found in the degree of potentiation among groups. In particular, increases of the CA1 response amplitudes were significantly lower in old rats impaired in spatial learning than in young at 30 (P < 0.05), 60, 90 and 120 min (P < 0.01) after restoring the normal Ca2+ concentration. On the contrary, no differences were observed between young animals and the old ones with good performances in spatial learning. The data suggest that amplitude of CA1 Ca(2+)-induced LTP in old rats is related to spatial learning abilities.

Selective reduction of hippocampal dentate frequency potentiation in aged rats with impaired place learning

Induction of posttetanic potentiation (PTP) and long-term potentiation (LTP) was analyzed in hippocampal slices obtained from a) young 6-month-old Sprague-Dawley (SD) rats, all of them performing well in the Morris Maze, and b) aged SD 20-month-old and Fischer 344 24-month-old rats showing different degrees of ability in the same test. After the application of an electrical tetanus 1 s, 100 Hz, 50 microA in the stratum radiatum, no significant differences were found in the percent of induction of both PTP and LTP in the CA1 area of hippocampal slices obtained from rats of different strains and ages. After the application of an electrical tetanus 1 s, 100 Hz, 50 microA in the stratum moleculare, a significant difference was found in the percent of dentate PTP induction in hippocampal slices obtained from rats of different ages. Specifically, dentate PTP induction was significantly (p < 0.01) higher in slices obtained from young SD rats, and from old SD rats with a better performance in the Morris maze, escape latency less than 10 s and 150 cm, than in slices obtained from old SD or Fischer 344 rats that had shown poor performance in the Morris Maze. On the contrary, no significant differences were found in the percent of dentate LTP in hippocampal slices obtained from rats of different strains and ages. The data demonstrate that the induction of hippocampal dentate high-frequency PTP is selectively reduced in old rats with impaired Morris Maze performance.

Behavioral and electrophysiological correlates of the quinolinic acid rat model of Huntington's disease in rats

The influence of bilateral intrastriatal injection of quinolinic acid (QA, 300 nmol) was studied in male Wistar rats. Behavioral and electrophysiological experiments were conducted in 15 lesioned plus 15 vehicle-injected (control) animals. With respect to control animals, QA-lesioned rats showed marked, statistically significant alterations from both the behavioral (greater motor activation in response to d-amphetamine, place-learning deficit in the Morris water maze), and the electroencephalographic (reduced voltage amplitude and EEG power at the level of frontal cortex) points of view. In addition, a significant loss in body weight and a marked striatal gliosis (GFAP staining) were observed in lesioned rats. Conversely, QA-lesioned rats did not show modifications in posttetanic potentiation (P.T.P.) or long-term potentiation (L.T.P.) in CA1 hippocampal area. The present results confirm that QA lesions of rat striatum may be regarded as a suitable model of Huntingtons disease (HD).

Different capability of N-methyl-D-aspartate antagonists to affect locomotor/exploratory activity of mice in a computerized on-line open field test

The effects of the competitive N-methyl-D-aspartate (NMDA) antagonists CGS 19755 and CPP, and of the noncompetitive NMDA antagonists PCP, MK 801, and dextromethorphan (DM) have been studied on the locomotor/exploratory activity of mice in a computerized on-line open field test. CGS 19755 (12.5-25 mg/kg, IP) induced a dose-dependent decrease in the locomotor/exploratory activity of mice; CPP (25-50 mg/kg, IP) did not present such an effect. PCP (1.25-10 mg/kg, IP) induced a dose-dependent increase/decrease in the locomotor/exploratory activity of mice, and DM (25-50 mg/kg, IP) and MK 801 (0.125-0.250 mg/kg, IP) increased it. The data show that NMDA antagonists affect locomotor/exploratory activity of mice in different ways, inducing both potentiating and inhibitory effects.

Does hypertension alone lead to cognitive decline in spontaneously hypertensive rats

Spontaneously hypertensive rats (SHR) represent an animal model of cognitive decline associated with hypertension. Few studies have systematically investigated this decline in aging. We assessed spatial learning performances of SHR, the genetically similar Wistar-Kyoto rats (WKY), and Sprague-Dawley rats (SD), in a water maze (WM) task. The following age ranges were studied: 6, 12, and 20 months. The results demonstrated that all three strains show the same level of impairment at the age of 20 months. The rate of cognitive decline, however, is different: both SHR and WKY show moderate degrees of impairment at all age ranges, while SD display good cognitive abilities at 6 months, declining at 12, and reaching the performance level of the other two groups at 20 months. Besides, the nine groups showed substantial differences in swim velocity and WKY exhibited a peculiar motor behavior. These results suggest the following: (a) the decline in cognitive level exhibits different trends in the three strains; (b) learning impairment of aged SHR might not be entirely explained by hypertension; (c) WKY should be used cautiously as normotensive control for SHR, due to their unusual behavior and low learning abilities; (d) analysis of escape distances is mandatory for the comparison of different strains in the WM test.

Behavioral effects of Rho GTPase modulation in a model of Alzheimer's disease

Small GTPases of the Rho family, including Rho, Rac and CDC42 subfamilies, play key role in neural connectivity and cognition. The pharmacological modulation of these regulatory proteins is associated with enhancement of learning and memory. We sought to determine whether the modulation of cerebral Rho GTPases may correct behavioral disturbances in a mouse model of Alzheimers disease (AD). TgCRND8 mice show early-onset Abeta amyloid deposits associated with deficits in several cognitive tasks. We report that four-month old TgCRND8 mice display (a) increased locomotor activity in an open field, (b) mild deficits in the learning of a fixed platform position in a water maze task. More markedly, after displacement of the escape platform, TgCRND8 mice exhibit impairment in the learning of the novel position (reversal learning), as they perseverate searching in the familiar position. The administration of the Rho GTPase activator Cytotoxic Necrotizing Factor 1 (CNF1, 1.0 fmol kg(-1) intracerebroventricularly) reduces locomotor hyperactivity and corrects the deficits in reversal learning, thus re-establishing normal behavioral plasticity. We conclude that the pharmacological modulation of Rho GTPase signaling might be beneficial for the treatment of AD. Reversal learning in TgCRND8 mice may represent a convenient pre-clinical assay for the efficacy of therapeutic interventions in AD.

The Rho GTPase activating CNF1 improves associative working memory for object-in-place

Cerebral Rho GTPases are crucially involved in cognitive abilities. This activity is thought to be related to the regulation of actin polymerization and, thereby, of the shape of the dendritic tree. Here we report that Cytotoxic Necrotizing Factor 1 (CNF1, 1fmol/kgicv), a bacterial protein endowed with Rho GTPase activating properties, enhances working memory for object location/discrimination in C57BL/6 mice. CNF1 selectively increased the exploration of a specific familiar object moved to a position that had been previously occupied by another familiar object. Conversely, the treatment left unaffected (i) exploration of a familiar object moved to a location that was previously unoccupied and (ii) exploration of a novel object. The effects were associated with changes in Rho GTPase status, since CNF1 C866S, a recombinant CNF1 in which the enzymatic activity was abolished through substitution of serine to cysteine at position 866, was ineffective in all the experiments. The study suggests that working memory for specific object-location associations critically depends on neural connectivity. It also confirms the therapeutic potential of the manipulation of Rho GTPase signaling in the modulation of memory processes.