Oligomeric amyloid-β (Aβ) 1-42 disrupts synaptic function at an early stage of Alzheimer's disease (AD). Multiple posttranslational modifications of Aβ have been identified, among which N-terminally truncated forms are the most abundant. It is not clear, however, whether modified species can induce synaptic dysfunction on their own and how altered biochemical properties can contribute to the synaptotoxic mechanisms. Here, we show that a prominent isoform, pyroglutamated Aβ3(pE)-42, induces synaptic dysfunction to a similar extent like Aβ1-42 but by clearly different mechanisms. In contrast to Aβ1-42, Aβ3(pE)-42 does not directly associate with synaptic membranes or the prion protein but is instead taken up by astrocytes and potently induces glial release of the proinflammatory cytokine TNFα. Moreover, Aβ3(pE)-42-induced synaptic dysfunction is not related to NMDAR signalling and Aβ3(pE)-42-induced impairment of synaptic plasticity cannot be rescued by D1-agonists. Collectively, the data point to a scenario where neuroinflammatory processes together with direct synaptotoxic effects are caused by posttranslational modification of soluble oligomeric Aβ and contribute synergistically to the onset of synaptic dysfunction in AD.
Highlights d Cav1 channels and RyRs form a complex with KCa3.1 channels in hippocampal neurons d The complex is maintained by junctophilin 3 and 4 proteins that tether ER-PM junctions d sAHP and spike accommodation in hippocampal neurons depends on JPH3 and JPH4 proteins
Ca1 L-type calcium channels are key to regulating neuronal excitability, with the range of functional roles enhanced by interactions with calmodulin, accessory proteins, or CaMKII that modulate channel activity. In hippocampal pyramidal cells, a prominent elevation of Ca1 activity is apparent in late channel openings that can last for seconds following a depolarizing stimulus train. The current study tested the hypothesis that a reported interaction among Ca1.3 channels, the scaffolding protein densin, and CaMKII could generate a facilitation of channel activity that outlasts a depolarizing stimulus. We found that Ca1.3 but not Ca1.2 channels exhibit a long-duration calcium-dependent facilitation (L-CDF) that lasts up to 8 s following a brief 50 Hz stimulus train, but only when coexpressed with densin and CaMKII. To test the physiological role for Ca1.3 L-CDF, we coexpressed the intermediate-conductance KCa3.1 potassium channel, revealing a strong functional coupling to Ca1.3 channel activity that was accentuated by densin and CaMKII. Moreover, the Ca1.3-densin-CaMKII interaction gave rise to an outward tail current of up to 8 s duration following a depolarizing stimulus in both tsA-201 cells and male rat CA1 pyramidal cells. A slow afterhyperpolarization in pyramidal cells was reduced by a selective block of Ca1 channels by isradipine, a CaMKII blocker, and siRNA knockdown of densin, and spike frequency increased upon selective block of Ca1 channel conductance. The results are important in revealing a Ca1.3-densin-CaMKII interaction that extends the contribution of Ca1.3 calcium influx to a time frame well beyond a brief input train. Ca1 L-type calcium channels play a key role in regulating the output of central neurons by providing calcium influx during repetitive inputs. This study identifies a long-duration calcium-dependent facilitation (L-CDF) of Ca1.3 channels that depends on the scaffolding protein densin and CaMKII and that outlasts a depolarizing stimulus by seconds. We further show a tight functional coupling between Ca1.3 calcium influx and the intermediate-conductance KCa3.1 potassium channel that promotes an outward tail current of up to 8 s following a depolarizing stimulus. Tests in CA1 hippocampal pyramidal cells reveal that a slow AHP is reduced by blocking different components of the Ca1.3-densin-CaMKII interaction, identifying an important role for Ca1.3 L-CDF in regulating neuronal excitability.
Stable expression of pannexin 1 (Panx1) and pannexin 3 (Panx3) resulted in functional gap junctions (GJs) in HeLa cells, but not in Neuro-2a (N2a) or PC-12 cells. The glycosylation pattern of expressed Panx1 varied greatly among different cell lines. In contrast to connexin (Cx) containing GJs (Cx-GJs), junctional conductance (Gj) of pannexin GJs (Panx-GJs) is very less sensitive to junctional voltage. Both Panx1 and Panx3 junctions favoured anionic dyes over cations to permeate. Though, carbenoxolone (CBX) and probenecid blocked Panx1 hemichannel activity, they had no effect on Panx1-GJs or Panx3-GJs. Extracellular loop 1 (E1) of Panx1 possibly bears the binding pocket. The Cx-GJ blocker heptanol blocked neither Panx1 hemichannel nor Panx-GJs. Unlike the GJs formed by most Cxs, CO2 did not uncouple Panx-GJs completely. Oxygen and glucose deprivation (OGD) caused lesser uncoupling of Panx-GJs compared to Cx43-GJs. These findings demonstrate properties of Panx-GJs that are distinctly different from Cx-GJs.
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