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Impaired ILK Function Is Associated with Deficits in Hippocampal Based Memory and Synaptic Plasticity in a FASD Rat Model.

Bhattacharya D, Dunaway EP, Bhattacharya S, Bloemer J, Buabeid M, Escobar M, Suppiramaniam V, Dhanasekaran M - PLoS ONE (2015)

Bottom Line: This reduced memory performance was consistent with decrease in LTP as compared to controls.Furthermore, immunoprecipitation of ILK revealed a decreased interaction with GluR2.These impairments appear to be mediated by reduced GSK3β regulation and increased synaptic stabilization of the calcium-impermeable GluR2 AMPA receptors.

View Article: PubMed Central - PubMed

Affiliation: Department of Drug, Discovery and Development, Auburn University, Auburn, Alabama, United States of America.

ABSTRACT
Fetal Alcohol Spectrum Disorder (FASD) is an umbrella term that encompasses a wide range of anatomical and behavioral problems in children who are exposed to alcohol during the prenatal period. There is no effective treatment for FASD, because of lack of complete characterization of the cellular and molecular mechanisms underlying this condition. Alcohol has been previously characterized to affect integrins and growth factor signaling receptors. Integrin Linked Kinase (ILK) is an effector of integrin and growth-factor signaling which regulates various signaling processes. In FASD, a downstream effector of ILK, Glycogen Synthase Kinase 3β (GSK3β) remains highly active (reduced Ser9 phosphorylation). GSK3β has been known to modulate glutamate receptor trafficking and channel properties. Therefore, we hypothesize that the cognitive deficits accompanying FASD are associated with impairments in the ILK signaling pathway. Pregnant Sprague Dawley rats consumed a "moderate" amount of alcohol throughout gestation, or a calorie-equivalent sucrose solution. Contextual fear conditioning was used to evaluate memory performance in 32-33-day-old pups. Synaptic plasticity was assessed in the Schaffer Collateral pathway, and hippocampal protein lysates were used to evaluate ILK signaling. Alcohol exposed pups showed impaired contextual fear conditioning, as compared to control pups. This reduced memory performance was consistent with decrease in LTP as compared to controls. Hippocampal ILK activity and GSK3β Ser21/9 phosphorylation were significantly lower in alcohol-exposed pups than controls. Increased synaptic expression of GluR2 AMPA receptors was observed with immunoprecipitation of post-synaptic density protein 95 (PSD95). Furthermore, immunoprecipitation of ILK revealed a decreased interaction with GluR2. The ILK pathway appears to play a significant role in memory and synaptic plasticity impairments in FASD rats. These impairments appear to be mediated by reduced GSK3β regulation and increased synaptic stabilization of the calcium-impermeable GluR2 AMPA receptors.

No MeSH data available.


Related in: MedlinePlus

Prenatal alcohol impairs hippocampal-based contextual fear memory.(a) four day lick suppression behavioral assay on control (n = 10) and alcohol groups (n = 16). Days 1 and 2 constituted training of the licking response in Context B, followed by conditioning in Context A on Day 3. A 0.65-mA scrambled footshock was delivered after 180th s of context exposure during the conditioning day. The latency to complete 50 licks in Context A were used as a measure of memory of the conditioning experience. (b) Time to complete 50 licks was calculated for the shock and no shock alcohol groups, compared to their nonexposed counterparts. (c) cumulative latencies to complete 50 licks are shown in 10 lick intervals for control and alcohol groups for shock and no shock category respectively. Differences were taken as statistically significant if p < 0.05.
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pone.0135700.g001: Prenatal alcohol impairs hippocampal-based contextual fear memory.(a) four day lick suppression behavioral assay on control (n = 10) and alcohol groups (n = 16). Days 1 and 2 constituted training of the licking response in Context B, followed by conditioning in Context A on Day 3. A 0.65-mA scrambled footshock was delivered after 180th s of context exposure during the conditioning day. The latency to complete 50 licks in Context A were used as a measure of memory of the conditioning experience. (b) Time to complete 50 licks was calculated for the shock and no shock alcohol groups, compared to their nonexposed counterparts. (c) cumulative latencies to complete 50 licks are shown in 10 lick intervals for control and alcohol groups for shock and no shock category respectively. Differences were taken as statistically significant if p < 0.05.

Mentions: Contextual fear conditioning is mainly dependent on hippocampal function. Dorsal hippocampus has a significant role in conditioned contextual freezing [23, 24]. Twenty-six animals served as subjects in this experiment. They were assigned to two groups, Alcohol (Eth, n = 16) or Control (Ctrl, n = 10). All animals were approximately 33 days old at the initiation of the behavioral portion of the study (average weight was 200 g). Each group was further randomly divided into shock and no shock subgroups. Animals were first trained to lick from a water spout in a novel environment, Context B. Conditioning was conducted in a different context, Context A, which was paired with a shock (Fig 1A). Memory of the association between Context A and the shock was assessed by placing the animals back in Context A, allowing them to begin licking from the spout, and recording the number of licks. The testing session was terminated at 10 min; thus, 600 s represent ceiling latency. Subjects’ access to water was gradually restricted to one hour per day over the week prior to initiation of the study, provided approximately one hour after completion of each day’s session. All conditioning procedures occurred in standard MedAssociates rat operant chambers and the experiments were scored by individuals who were blinded to the condition. Two modifications of these chambers served to create Context A (chambers with no additional cues), and Context B (a different set of chambers, modified using a striped pattern to cover the otherwise clear walls). Behavioral training and assessment occurred over 4 days, as follows. On Days 1 and 2 (lick training), subjects were trained to lick for water in Context B during two 30-min sessions. On Day 3 (conditioning), subjects were placed in Context A for 270 s. A 0.65-mA electric foot shock was delivered at 180 s for a duration of 2 s. Water bottles were not available during the conditioning session. On Day 4 (memory assessment), subjects were exposed to Context A for a 10 min period with water bottles available; this test evaluated recall of the association between Context A and the foot shock. Timing at which each lick was produced was recorded. Rats ‘freeze’ in anticipation of shock; thus, longer latencies to drink were assumed to reflect higher expectation of shock to be delivered in Context A (i.e., stronger memory of the conditioning experience). Latencies are skewed to the right; thus, a log (base 10) transformation was applied to the data to meet the normality criterion of parametric statistics. A 2 (treatment: alcohol vs. control) x 2 (condition: shock vs. no shock) analysis of variance (ANOVA) was conducted on the latency to complete 50 licks, assessed from the moment the first lick was produced until the 50th lick was produced.


Impaired ILK Function Is Associated with Deficits in Hippocampal Based Memory and Synaptic Plasticity in a FASD Rat Model.

Bhattacharya D, Dunaway EP, Bhattacharya S, Bloemer J, Buabeid M, Escobar M, Suppiramaniam V, Dhanasekaran M - PLoS ONE (2015)

Prenatal alcohol impairs hippocampal-based contextual fear memory.(a) four day lick suppression behavioral assay on control (n = 10) and alcohol groups (n = 16). Days 1 and 2 constituted training of the licking response in Context B, followed by conditioning in Context A on Day 3. A 0.65-mA scrambled footshock was delivered after 180th s of context exposure during the conditioning day. The latency to complete 50 licks in Context A were used as a measure of memory of the conditioning experience. (b) Time to complete 50 licks was calculated for the shock and no shock alcohol groups, compared to their nonexposed counterparts. (c) cumulative latencies to complete 50 licks are shown in 10 lick intervals for control and alcohol groups for shock and no shock category respectively. Differences were taken as statistically significant if p < 0.05.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4549293&req=5

pone.0135700.g001: Prenatal alcohol impairs hippocampal-based contextual fear memory.(a) four day lick suppression behavioral assay on control (n = 10) and alcohol groups (n = 16). Days 1 and 2 constituted training of the licking response in Context B, followed by conditioning in Context A on Day 3. A 0.65-mA scrambled footshock was delivered after 180th s of context exposure during the conditioning day. The latency to complete 50 licks in Context A were used as a measure of memory of the conditioning experience. (b) Time to complete 50 licks was calculated for the shock and no shock alcohol groups, compared to their nonexposed counterparts. (c) cumulative latencies to complete 50 licks are shown in 10 lick intervals for control and alcohol groups for shock and no shock category respectively. Differences were taken as statistically significant if p < 0.05.
Mentions: Contextual fear conditioning is mainly dependent on hippocampal function. Dorsal hippocampus has a significant role in conditioned contextual freezing [23, 24]. Twenty-six animals served as subjects in this experiment. They were assigned to two groups, Alcohol (Eth, n = 16) or Control (Ctrl, n = 10). All animals were approximately 33 days old at the initiation of the behavioral portion of the study (average weight was 200 g). Each group was further randomly divided into shock and no shock subgroups. Animals were first trained to lick from a water spout in a novel environment, Context B. Conditioning was conducted in a different context, Context A, which was paired with a shock (Fig 1A). Memory of the association between Context A and the shock was assessed by placing the animals back in Context A, allowing them to begin licking from the spout, and recording the number of licks. The testing session was terminated at 10 min; thus, 600 s represent ceiling latency. Subjects’ access to water was gradually restricted to one hour per day over the week prior to initiation of the study, provided approximately one hour after completion of each day’s session. All conditioning procedures occurred in standard MedAssociates rat operant chambers and the experiments were scored by individuals who were blinded to the condition. Two modifications of these chambers served to create Context A (chambers with no additional cues), and Context B (a different set of chambers, modified using a striped pattern to cover the otherwise clear walls). Behavioral training and assessment occurred over 4 days, as follows. On Days 1 and 2 (lick training), subjects were trained to lick for water in Context B during two 30-min sessions. On Day 3 (conditioning), subjects were placed in Context A for 270 s. A 0.65-mA electric foot shock was delivered at 180 s for a duration of 2 s. Water bottles were not available during the conditioning session. On Day 4 (memory assessment), subjects were exposed to Context A for a 10 min period with water bottles available; this test evaluated recall of the association between Context A and the foot shock. Timing at which each lick was produced was recorded. Rats ‘freeze’ in anticipation of shock; thus, longer latencies to drink were assumed to reflect higher expectation of shock to be delivered in Context A (i.e., stronger memory of the conditioning experience). Latencies are skewed to the right; thus, a log (base 10) transformation was applied to the data to meet the normality criterion of parametric statistics. A 2 (treatment: alcohol vs. control) x 2 (condition: shock vs. no shock) analysis of variance (ANOVA) was conducted on the latency to complete 50 licks, assessed from the moment the first lick was produced until the 50th lick was produced.

Bottom Line: This reduced memory performance was consistent with decrease in LTP as compared to controls.Furthermore, immunoprecipitation of ILK revealed a decreased interaction with GluR2.These impairments appear to be mediated by reduced GSK3β regulation and increased synaptic stabilization of the calcium-impermeable GluR2 AMPA receptors.

View Article: PubMed Central - PubMed

Affiliation: Department of Drug, Discovery and Development, Auburn University, Auburn, Alabama, United States of America.

ABSTRACT
Fetal Alcohol Spectrum Disorder (FASD) is an umbrella term that encompasses a wide range of anatomical and behavioral problems in children who are exposed to alcohol during the prenatal period. There is no effective treatment for FASD, because of lack of complete characterization of the cellular and molecular mechanisms underlying this condition. Alcohol has been previously characterized to affect integrins and growth factor signaling receptors. Integrin Linked Kinase (ILK) is an effector of integrin and growth-factor signaling which regulates various signaling processes. In FASD, a downstream effector of ILK, Glycogen Synthase Kinase 3β (GSK3β) remains highly active (reduced Ser9 phosphorylation). GSK3β has been known to modulate glutamate receptor trafficking and channel properties. Therefore, we hypothesize that the cognitive deficits accompanying FASD are associated with impairments in the ILK signaling pathway. Pregnant Sprague Dawley rats consumed a "moderate" amount of alcohol throughout gestation, or a calorie-equivalent sucrose solution. Contextual fear conditioning was used to evaluate memory performance in 32-33-day-old pups. Synaptic plasticity was assessed in the Schaffer Collateral pathway, and hippocampal protein lysates were used to evaluate ILK signaling. Alcohol exposed pups showed impaired contextual fear conditioning, as compared to control pups. This reduced memory performance was consistent with decrease in LTP as compared to controls. Hippocampal ILK activity and GSK3β Ser21/9 phosphorylation were significantly lower in alcohol-exposed pups than controls. Increased synaptic expression of GluR2 AMPA receptors was observed with immunoprecipitation of post-synaptic density protein 95 (PSD95). Furthermore, immunoprecipitation of ILK revealed a decreased interaction with GluR2. The ILK pathway appears to play a significant role in memory and synaptic plasticity impairments in FASD rats. These impairments appear to be mediated by reduced GSK3β regulation and increased synaptic stabilization of the calcium-impermeable GluR2 AMPA receptors.

No MeSH data available.


Related in: MedlinePlus