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Prolonged Abeta treatment leads to impairment in the ability of primary cortical neurons to maintain K+ and Ca2+ homeostasis.

Shabala L, Howells C, West AK, Chung RS - Mol Neurodegener (2010)

Bottom Line: This decrease in survival correlated with increased K+ efflux from the cells.Ca2+ uptake was significantly higher only after prolonged Abeta treatment with 2.5-fold increase in total Ca2+ uptake over 20 min post glutamate application after six days of Abeta treatment or longer (P < 0.05).Our data suggest that long term exposure to Abeta is detrimental because it reduces the ability of cortical neurons to maintain K+ and Ca2+ homeostasis in response to glutamate challenge, a response that might underlie the early symptoms of AD.

View Article: PubMed Central - HTML - PubMed

Affiliation: NeuroRepair Group, Menzies Research Institute, University of Tasmania, Private Bag 23, Hobart, Tasmania, 7001, Australia. L.Shabala@utas.edu.au.

ABSTRACT

Background: Alzheimer's disease (AD) is a progressive neurodegenerative disease, characterised by the formation of insoluble amyloidogenic plaques and neurofibrillary tangles. Beta amyloid (Abeta) peptide is one of the main constituents in Abeta plaques, and is thought to be a primary causative agent in AD. Neurons are likely to be exposed to chronic, sublethal doses of Abeta over an extended time during the pathogenesis of AD, however most studies published to date using in vitro models have focussed on acute studies. To experimentally model the progressive pathogenesis of AD, we exposed primary cortical neurons daily to 1 muM of Abeta1-40 over 7 days and compared their survival with age-similar untreated cells. We also investigated whether chronic Abeta exposure affects neuronal susceptibility to the subsequent acute excitotoxicity induced by 10 muM glutamate and assessed how Ca2+ and K+ homeostasis were affected by either treatment.

Results: We show that continuous exposure to 1 muM Abeta1-40 for seven days decreased survival of cultured cortical neurons by 20%. This decrease in survival correlated with increased K+ efflux from the cells. One day treatment with 1 muM Abeta followed by glutamate led to a substantially higher K+ efflux than in the age-similar untreated control. This difference further increased with the duration of the treatment. K+ efflux also remained higher in Abeta treated cells 20 min after glutamate application leading to 2.8-fold higher total K+ effluxed from the cells compared to controls. Ca2+ uptake was significantly higher only after prolonged Abeta treatment with 2.5-fold increase in total Ca2+ uptake over 20 min post glutamate application after six days of Abeta treatment or longer (P < 0.05).

Conclusions: Our data suggest that long term exposure to Abeta is detrimental because it reduces the ability of cortical neurons to maintain K+ and Ca2+ homeostasis in response to glutamate challenge, a response that might underlie the early symptoms of AD. The observed inability to maintain K+ homeostasis might furthermore be useful in future studies as an early indicator of pathological changes in response to Abeta.

No MeSH data available.


Related in: MedlinePlus

Kinetics of Ca2+ fluxes in response to acute application of 10 μM glutamate. Ca2+ fluxes were measured from age-similar control cells (A) and primary cortical neurons treated daily with 1 μM Aβ1-40 for six days (B). Flux values were recorded for 5 min before glutamate application (-5 to 0 min) and 25 min afterwards with data acquired at a rate of 10 samples/sec and averaged over every 6 sec. Glutamate (10 μM) was applied at zero time as indicated by an arrow and led to Ca2+ influx into cultured cortical neurons (positive flux values). One (out of 7) typical example is shown.
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Figure 5: Kinetics of Ca2+ fluxes in response to acute application of 10 μM glutamate. Ca2+ fluxes were measured from age-similar control cells (A) and primary cortical neurons treated daily with 1 μM Aβ1-40 for six days (B). Flux values were recorded for 5 min before glutamate application (-5 to 0 min) and 25 min afterwards with data acquired at a rate of 10 samples/sec and averaged over every 6 sec. Glutamate (10 μM) was applied at zero time as indicated by an arrow and led to Ca2+ influx into cultured cortical neurons (positive flux values). One (out of 7) typical example is shown.

Mentions: We next investigated whether neurons exposed to prolonged Aβ1-40 treatment had increased susceptibility to subsequent excitotoxicity induced by 200 μM glutamate. While glutamate treatment for 24 hours killed approximately 20% of neurons in the absence of any Aβ1-40 treatment (Figure 3A), we found that seven days of pre-treatment with Aβ1-40 did not further elevate neuronal susceptibility to glutamate-induced neurotoxicity (Figure 3B). We then examined K+ and Ca2+ fluxes in neurons which were treated with 1 μM Aβ1-40 daily for a period from one to eight days and then challenged with 10 μM of glutamate. Using MIFE, net fluxes of K+ and Ca2+ were monitored continuously throughout the experiment starting before the glutamate challenge and up to 30 min afterwards with data acquired every 6 sec. Typical examples of kinetics of net K+ and Ca2+ fluxes in response to challenge with glutamate are shown in Figures 4A-B and 5A-B, respectively.


Prolonged Abeta treatment leads to impairment in the ability of primary cortical neurons to maintain K+ and Ca2+ homeostasis.

Shabala L, Howells C, West AK, Chung RS - Mol Neurodegener (2010)

Kinetics of Ca2+ fluxes in response to acute application of 10 μM glutamate. Ca2+ fluxes were measured from age-similar control cells (A) and primary cortical neurons treated daily with 1 μM Aβ1-40 for six days (B). Flux values were recorded for 5 min before glutamate application (-5 to 0 min) and 25 min afterwards with data acquired at a rate of 10 samples/sec and averaged over every 6 sec. Glutamate (10 μM) was applied at zero time as indicated by an arrow and led to Ca2+ influx into cultured cortical neurons (positive flux values). One (out of 7) typical example is shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2927593&req=5

Figure 5: Kinetics of Ca2+ fluxes in response to acute application of 10 μM glutamate. Ca2+ fluxes were measured from age-similar control cells (A) and primary cortical neurons treated daily with 1 μM Aβ1-40 for six days (B). Flux values were recorded for 5 min before glutamate application (-5 to 0 min) and 25 min afterwards with data acquired at a rate of 10 samples/sec and averaged over every 6 sec. Glutamate (10 μM) was applied at zero time as indicated by an arrow and led to Ca2+ influx into cultured cortical neurons (positive flux values). One (out of 7) typical example is shown.
Mentions: We next investigated whether neurons exposed to prolonged Aβ1-40 treatment had increased susceptibility to subsequent excitotoxicity induced by 200 μM glutamate. While glutamate treatment for 24 hours killed approximately 20% of neurons in the absence of any Aβ1-40 treatment (Figure 3A), we found that seven days of pre-treatment with Aβ1-40 did not further elevate neuronal susceptibility to glutamate-induced neurotoxicity (Figure 3B). We then examined K+ and Ca2+ fluxes in neurons which were treated with 1 μM Aβ1-40 daily for a period from one to eight days and then challenged with 10 μM of glutamate. Using MIFE, net fluxes of K+ and Ca2+ were monitored continuously throughout the experiment starting before the glutamate challenge and up to 30 min afterwards with data acquired every 6 sec. Typical examples of kinetics of net K+ and Ca2+ fluxes in response to challenge with glutamate are shown in Figures 4A-B and 5A-B, respectively.

Bottom Line: This decrease in survival correlated with increased K+ efflux from the cells.Ca2+ uptake was significantly higher only after prolonged Abeta treatment with 2.5-fold increase in total Ca2+ uptake over 20 min post glutamate application after six days of Abeta treatment or longer (P < 0.05).Our data suggest that long term exposure to Abeta is detrimental because it reduces the ability of cortical neurons to maintain K+ and Ca2+ homeostasis in response to glutamate challenge, a response that might underlie the early symptoms of AD.

View Article: PubMed Central - HTML - PubMed

Affiliation: NeuroRepair Group, Menzies Research Institute, University of Tasmania, Private Bag 23, Hobart, Tasmania, 7001, Australia. L.Shabala@utas.edu.au.

ABSTRACT

Background: Alzheimer's disease (AD) is a progressive neurodegenerative disease, characterised by the formation of insoluble amyloidogenic plaques and neurofibrillary tangles. Beta amyloid (Abeta) peptide is one of the main constituents in Abeta plaques, and is thought to be a primary causative agent in AD. Neurons are likely to be exposed to chronic, sublethal doses of Abeta over an extended time during the pathogenesis of AD, however most studies published to date using in vitro models have focussed on acute studies. To experimentally model the progressive pathogenesis of AD, we exposed primary cortical neurons daily to 1 muM of Abeta1-40 over 7 days and compared their survival with age-similar untreated cells. We also investigated whether chronic Abeta exposure affects neuronal susceptibility to the subsequent acute excitotoxicity induced by 10 muM glutamate and assessed how Ca2+ and K+ homeostasis were affected by either treatment.

Results: We show that continuous exposure to 1 muM Abeta1-40 for seven days decreased survival of cultured cortical neurons by 20%. This decrease in survival correlated with increased K+ efflux from the cells. One day treatment with 1 muM Abeta followed by glutamate led to a substantially higher K+ efflux than in the age-similar untreated control. This difference further increased with the duration of the treatment. K+ efflux also remained higher in Abeta treated cells 20 min after glutamate application leading to 2.8-fold higher total K+ effluxed from the cells compared to controls. Ca2+ uptake was significantly higher only after prolonged Abeta treatment with 2.5-fold increase in total Ca2+ uptake over 20 min post glutamate application after six days of Abeta treatment or longer (P < 0.05).

Conclusions: Our data suggest that long term exposure to Abeta is detrimental because it reduces the ability of cortical neurons to maintain K+ and Ca2+ homeostasis in response to glutamate challenge, a response that might underlie the early symptoms of AD. The observed inability to maintain K+ homeostasis might furthermore be useful in future studies as an early indicator of pathological changes in response to Abeta.

No MeSH data available.


Related in: MedlinePlus