A.C.L. Gianlorenço

Microinjection of histamine into the cerebellar vermis impairs emotional memory consolidation in mice.

The biogenic amine histamine is an important neurotransmitter in the central nervous system that has been implicated in learning and memory processes. Experimental evidence indicates that the role of the cerebellum may be more complex than the simple regulation of motor responses, and recent studies have demonstrated significant involvement of the cerebellum in emotional memory consolidation. This study investigated the effect of histamine microinjected into the cerebellar vermis on emotional memory consolidation in mice in the elevated plus-maze (EPM). The cerebellar vermis of male mice (Swiss Albino) were implanted with guide cannulae. The mice weighed between 25 and 30 g. After three days of recovery, behavioral tests in the EPM were performed on two consecutive days; the testing periods were called, Trial 1 and Trial 2. Immediately after Trial 1, the animals received microinjections of histamine in the cerebellar vermis (0.54, 1.36, 2.72, and 4.07 nmol/0.1 μl). On both days, the test sessions were recorded to enable analysis of behavioral measures. The decrease in open arm exploration (% entries and % time spent in the open arms) in Trial 2 relative to Trial 1 was used as a measure of learning and memory. The data were analyzed using One-way Analysis of Variance (ANOVA) and Duncans tests. The percentage of open arm entries (%OAE) and the percentage of time spent in the open arms (%OAT) were reduced in Trial 2 relative to Trial 1 for the control group; the same was true for the group that was microinjected with histamine at doses of 0.54 (%OAE and %OAT) and 1.36 nmol (%OAT). However, when the animals received histamine at doses of 2.72 and 4.07 nmol, their open arm exploration did not decrease. No significant changes were observed in the number of enclosed arm entries (EAE), an EPM index of general exploratory activity. These results suggest that there is a dose-dependent inhibitory effect of histamine microinjected into the cerebellar vermis on emotional memory consolidation.

Emotional memory consolidation impairment induced by histamine is mediated by H1 but not H2 receptors

Histaminergic fibers are present in the molecular and granular layers of the cerebellum and have high density in the vermis and flocculus. Evidence indicates that the cerebellar vermis is involved in memory consolidation. Recently, we demonstrated that when histamine is microinjected into the cerebellar vermis it results in impaired emotional memory consolidation in mice that are submitted to the elevated plus maze (EPM). This study investigated whether histamine impairment was mediated by the H(1) or H(2) receptors. The cerebellar vermis of male mice (Swiss Albino) were implanted using a guide cannula. Three days after recovery, behavioral tests were performed in the EPM on two consecutive days (Trial 1 and Trial 2). Immediately after exposure to the EPM (Trial 1), animals received a microinjection of histaminergic drugs. In Experiment 1, saline (SAL) or histamine (HA, 4.07 nmol/0.1 μl) was microinjected 5 min after pretreatment with the H(1) antagonist chlorpheniramine (CPA, 0.16 nmol/0.1μl) or SAL. In Experiment 2, SAL or HA was microinjected into the mice 5 min after pretreatment with the H(2) antagonist ranitidine (RA, 2.85 nmol/0.1 μl) or SAL. Twenty-four hours later, the mice were re-exposed to the EPM (Trial 2) under the same experimental conditions but did not receive an injection. On both days, the test sessions were recorded to enable analysis of the behavioral measures. The decrease in open arm exploration (% entries and % time spent in the open arms) in Trial 2 relative to Trial 1 was used as a measure of learning and memory. The data were analyzed using the two-way analysis of variance (ANOVA) and Duncans tests. In Experiment 1, the Duncans test indicated that the mice entered the open arms less often (%OAE) and spent less time in the open arms (%OAT) in Trial 2 after being microinjected with SAL+SAL, SAL+CPA and CPA+HA. However, the animals that received SAL+HA did not enter the open arms less frequently or spend less time in them, which was significantly different from the CPA+HA group. The results of Experiment 2 demonstrated that the %OAE and %OAT in Trial 2 were different from Trial 1 for the groups that were microinjected with SAL+SAL and SAL+RA. The animals that were microinjected with RA+HA or with SAL+HA did not show a reduction in %OAE. These results demonstrate that the animals treated with HA did not avoid the open arms less on retesting and indicated that CPA did not alter the behavior parameters but did revert the histamine-induced impairment of memory consolidation. Furthermore, the H(2) antagonist RA, at the dose used in this study, did not affect memory consolidation and failed to revert histamine-induced impairment.

Intra-cerebellar microinjection of histamine enhances memory consolidation of inhibitory avoidance learning in mice via H2 receptors

Studies have demonstrated the relationship between the histaminergic system and the cerebellum, and we intend to investigate the role of the cerebellar histaminergic system on memory consolidation. This study investigated the effect of intra-cerebellar microinjection of histamine on memory retention of inhibitory avoidance in mice, and the role of H1 and H2 receptors in it. The cerebellar vermis of male mice were implanted with guide cannulae, and after three days of recovery, the inhibitory avoidance test was performed. Immediately after a training session, animals received a microinjection of histaminergic drugs: in the experiment 1, saline (SAL) or histamine (HA 0.54, 1.36, 2.72 or 4.07 nmol); experiment 2, SAL or 1.36 nmol HA 5 min after a pretreatment with 0.16 nmol chlorpheniramine (CPA) or SAL; and experiment 3, SAL or 1.36 nmol HA 5 min after a pretreatment with 2.85 nmol ranitidine (RA) or SAL. Twenty-four hours later, a retention test was performed. The data were analyzed using one-way analysis of variance (ANOVA) and Duncans tests. In experiment 1, animals microinjected with 1.36 nmol HA showed a higher latency to cross to the dark compartment compared to controls and to 2.72 and 4.07 nmol HA groups. In experiment 2, the combined infusions revealed difference between control (SAL+SAL) and SAL+HA and CPA+HA; while in the experiment 3 the analysis indicated differences in retention latency between mice injected with SAL+SAL and SAL+HA. The groups that received the H2 antagonist RA did not show difference compared to control. These results indicate that 1.36 nmol HA enhances memory consolidation of inhibitory avoidance learning in mice and that the pretreatment with H2 antagonist RA was able to prevent this effect.

l-histidine induces state-dependent memory deficit in mice mediated by H1 receptor

This study investigated the role of H(1) receptor in the state-dependent memory deficit induced by l-histidine (LH) in mice using Trial 1/2 protocol in the elevated plus-maze (EPM). The test was performed for two consecutive days: Trial 1 (T1) and Trial 2 (T2). Before both trials, mice received a combined injection i.p. of saline+saline (SAL/SAL), 500 mg/kg L-histidine+saline (LH/SAL), 500 mg/kg L-histidine+16 mg/kg chlorpheniramine (LH/CPA) or saline+16 mg/kg chlorpheniramine (SAL/CPA). The trials were performed in the EPM 10 min after the last injection. Each animal was placed in the center of the maze facing the open arm and had five minutes to explore it. On both days, test sessions were videotaped. The behavioral measures were scored from videotape. Data were analyzed based on Analysis of Variance (ANOVA) and the Fishers LSD test. The data showed no effects on anxiety since there was no difference between the SAL/SAL and the other groups in Trial 1, respectively, open arm entries (OAE), open arm time (OAT) and their percentages (%OAE and %OAT). During Trial 2, OAE, OAT, %OAE and %OAT were reduced in mice treated with SAL/SAL, LH/CPA and SAL/CPA, while the group LH/SAL did not show any difference in these measures. No significant changes were observed in enclosed arm entries (EAE), an EPM index of general exploratory activity. Thus, it can be suggested that LH induces emotional memory deficit and the treatment with chlorpheniramine was able to revert this effect, suggesting this action of LH was mediated by the H(1) receptor.

H1-histamine receptors in the amygdala are involved in emotional memory but do not mediate anxiety-related behaviors in mice submitted to EPM testing

This study investigated the role of amygdala H(1) receptors in state-dependent memory deficits induced by l-histidine (LH). Tests using an elevated plus-maze (EPM) were performed on two consecutive days: Trial 1 (T1) and Trial 2 (T2). Before each trial, mice were intraperitoneally (IP) injected with LH (500mg/kg). Two hours later, they were microinjected with the H(1) receptor antagonist, chlorpheniramine (CPA 0.016, 0.052, or 0.16 nmol/0.1μl), or saline (SAL) into the amygdala and submitted to the EPM. LH-CPA did not affect trial 1 performances in the EPM, which indicated that these drugs did not affect anxiety. Emotional memory, as revealed by a reduction in open arm exploration between both trials, was present in the SAL-SAL groups as well as in the SAL-CPA groups for the lower doses of CPA (0.016 and 0.052nmol). On the contrary, neither the LH-SAL group nor the LH-CPA groups exhibited this decrease in open arm activity between both trials, which reveals that LH impaired emotional memory. While intra-amygdalar CPA did not interact with LH effect, it impaired per se the emotional memory performances at the highest dose (0.16nmol). No significant changes were observed in the number of enclosed arm entries (EAE), an EPM index of general exploratory activity. These results may be attributed to a combined effect in the different nucleus of the amygdala. Taken together, these results suggest that the H(1) receptors in the amygdala are not implicated in anxiety-like behaviors but are involved in emotional states induced by the T1/T2 EPM protocol in mice.

Cerebellar vermis H2 receptors mediate fear memory consolidation in mice

Histaminergic fibers are present in the molecular and granular layers of the cerebellum and have a high density in the vermis and flocullus. Evidence supports that the cerebellar histaminergic system is involved in memory consolidation. Our recent study showed that histamine injections facilitate the retention of an inhibitory avoidance task, which was abolished by pretreatment with an H2 receptor antagonist. In the present study, we investigated the effects of intracerebellar post training injections of H1 and H2 receptor antagonists as well as the selective H2 receptor agonist on fear memory consolidation. The cerebellar vermi of male mice were implanted with guide cannulae, and after three days of recovery, the inhibitory avoidance test was performed. Immediately after a training session, animals received a microinjection of the following histaminergic drugs: experiment 1, saline or chlorpheniramine (0.016, 0.052 or 0.16 nmol); experiment 2, saline or ranitidine (0.57, 2.85 or 5.07 nmol); and experiment 3, saline or dimaprit (1, 2 or 4 nmol). Twenty-four hours later, a retention test was performed. The data were analyzed using one-way analysis of variance (ANOVA) and Duncans tests. Animals microinjected with chlorpheniramine did not show any behavioral effects at the doses that we used. Intra-cerebellar injection of the H2 receptor antagonist ranitidine inhibited, while the selective H2 receptor agonist dimaprit facilitated, memory consolidation, suggesting that H2 receptors mediate memory consolidation in the inhibitory avoidance task in mice.

Intracerebellar vermis histamine facilitates memory consolidation in the elevated T maze model

Experimental evidence suggests that the cerebellum plays a more complex role in learning than simply regulating the motor response. Rather, it is thought to play a significant role in the consolidation of emotional memory in mice. Due to the difficulty of interpreting fear and anxiety behaviors-the standard methodology for the study of the histaminergic system and emotional memory-in mice, we propose a behavioral assessment of mice subjected to the Elevated T-maze after histamine microinjection of the cerebellar vermis. Young male Swiss albino mice weighing 25-35g were used. In addition, locomotor activity was tested in an open field test. Our data suggest that histamine did not affect memory consolidation during escape or open field behavior at the doses used in this study. However, we observed a significant increase in inhibitory avoidance on the second day in animals receiving a dose of 6.8nmol/0.5μl, suggesting that histamine facilitates the consolidation of inhibitory avoidance in mice.

Effect of histamine H1 and H2 receptor antagonists, microinjected into cerebellar vermis, on emotional memory consolidation in mice

This study investigated the effects of histamine H1 or H2 receptor antagonists on emotional memory consolidation in mice submitted to the elevated plus maze (EPM). The cerebellar vermis of male mice (Swiss albino) was implanted using a cannula guide. Three days after recovery, behavioral tests were performed in the EPM on 2 consecutive days (T1 and T2). Immediately after exposure to the EPM (T1), animals received a microinjection of saline (SAL) or the H1 antagonist chlorpheniramine (CPA; 0.016, 0.052, or 0.16 nmol/0.1 µL) in Experiment 1, and SAL or the H2 antagonist ranitidine (RA; 0.57, 2.85, or 5.7 nmol/0.1 µL) in Experiment 2. Twenty-four hours later, mice were reexposed to the EPM (T2) under the same experimental conditions but they did not receive any injection. Data were analyzed using one-way ANOVA and the Duncan test. In Experiment 1, mice microinjected with SAL and with CPA entered the open arms less often (%OAE) and spent less time in the open arms (%OAT) in T2, and there was no difference among groups. The results of Experiment 2 demonstrated that the values of %OAE and %OAT in T2 were lower compared to T1 for the groups that were microinjected with SAL and 2.85 nmol/0.1 µL RA. However, when animals were microinjected with 5.7 nmol/0.1 µL RA, they did not show a reduction in %OAE and %OAT. These results demonstrate that CPA did not affect behavior at the doses used in this study, while 5.7 nmol/0.1 µL RA induced impairment of memory consolidation in the EPM.

Cholinergic agonist reverses H1-induced memory deficit in mice.

ABSTRACT This study investigated the effects of bilateral intraamygdalar microinjections of PNU‐282987, a nicotinic cholinergic agonist, on anxiety and the reversal of amnesia induced by chlorpheniramine (CPA), an H1 histaminergic antagonist, in mice subjected to the elevated plusmaze (EPM). Two experiments were performed with seventy‐nine adult male Swiss mice. The isolated microinjections of PNU‐282987 did not produce effects on emotional memory; however, the combined microinjections of PNU‐282987 and CPA were able to reverse the deficit in memory induced by CPA (ANOVA, p < 0.05). Taken together, these results suggest that intraamygdalar injections of PNU‐282987 did not induce effects on anxiety and emotional memory per se; however, concurrent microinjections of PNU‐282987 and CPA‐reverse amnesia induced‐CPA which is suggestive of an interaction between the histaminergic and cholinergic systems in the modulation of emotion memory acquisition in mice. HIGHLIGHTSChlorpheniramine and PNU‐282987 did not induce effects on anxiety in elevated plus maze.The PNU‐282987 per se did not produce effects on emotional memory.Microinjections of PNU‐282987 reverse memory deficits induced by chlorpheniramine.

Intra-vermis H4 receptor agonist impairs performance in anxiety- and fear-mediated models

The neural histaminergic system modulates cognitive performance in various animal models. However, little is known about the effects of the H4 histaminergic receptor in the central nervous system. The purpose of this study was to investigate the effect of histaminergic H4 agonist VUF-8430 microinjection into the cerebellar vermis on the consolidation of emotional memory in mice subjected to the elevated plus maze (EPM) and inhibitory avoidance task (IAT). All experiments were performed on two consecutive days: exposure (T1 and D1) and 24h after, which we called re-exposure (T2 and D2). The animals received saline (SAL) or VUF (0.15 nmol; 0.49 nmol; 1.48 nmol/0.1μl) administered post-exposure. Experiment 1 was conducted in the EPM, and the animals were free to explore the maze for 5min. In T1, immediately after exposure, the pharmacological treatment was given; in T2, there was only re-exposure to the EPM. Experiment 2 involved the IAT, and the pharmacological treatment was provided post-D1; in D2, the animals were only re-exposed to the IAT. In Experiment 1, increased open arm exploration (% open arm entries and% open arms time) for 0.49 and 1.48nmol of VUF were recorded in T2 compared to T1. In Experiment 2, a significant decrease in consolidation latency was recorded for the group that received 1.48nmol of VUF compared to the SAL group in D2. These results indicate that a 1.48nmol VUF microinjection into the cerebellar vermis impaired performance in both models, even though one model was anxiety-mediated (EPM) and the other was fear-mediated (IAT).