The actions of beta amyloid on human microglia involve Fc receptor and K+ channel activation

Sonia Franciosi, Han S. Jeong, Hyun B. Choi, James G. McLarnon, Department of Pharmacology & Therapeutics, University of British Columbia, Vancouver, Canada.

Stimulation of microglia, the immune cells of the central nervous system, with the Alzheimer’s plaque protein, beta amyloid (Aβ), results in the release of a plethora of inflammatory mediators and neurotoxic products. Little is known about the mechanisms involved in Aβ actions on microglia. The effects of Aβ1-42 (5 µM) on electrophysiological properties and ion channel expression in cultured human microglia were investigated using whole-cell patch clamp and molecular biology techniques.

Current clamp experiments showed that acute Aβ1-42 induced an immediate transient depolarization (mean ± SEM, 8.9 ± 0.9 mV; n=5 cells) followed by a small hyperpolarization of microglia. Aβ42-1 and vehicle controls had no effect on membrane potential. The involvement of the Fc receptor in mediating cell depolarization was examined since this receptor is coupled to a non-selective cationic channel. Results using an antibody-binding assay showed that Aβ1-42 stimulation of microglia significantly increased Fc receptor expression from basal levels by 20 ± 3.9% at 30 min (n=6; p < 0.01) and by 36 ± 1.3% at 48 hrs incubation (n=6; p < 0.001). Inhibition of the Fc receptor abrogated the acute Aβ1-42 induced depolarization (n = 7). This result was confirmed in voltage clamp studies showing the acute effect of Aβ1-42 to induce an inward current (mean amplitude: 350 ± 10 pA; n=4) which was blocked by Fc receptor inhibition. Statistical significance was determined using a Student’s t-test with p < 0.05 considered significant.

In voltage clamp studies, acute application of Aβ1-42 also induced a non-inactivating outward K+ current (IK) in response to a depolarizing step from –60 mV to +20 mV (mean amplitude of 850 ± 102 pA; n=7); control currents in human microglia were minimal (mean amplitude 59 ± 6 pA; n=7). The rapid induction of IK with Aβ1-42 indicated that the current was either constitutively present in the membrane but non-functional or rapidly inserted into the membrane from intracellular pools. This current was likely mediated through a G protein since intracellular application of GTPγS (10 µM) also induced a non-inactivating outward K+ channel (mean amplitude of 1149 ± 113 pA; n=9) from control (mean amplitude 147 ± 49 pA; n=5) similar in properties to IK. Threshold for activation of the GTPγS induced outward K+ current (-28 ± 1.7 mV; n=4) and IK (-33.7 ± 2.4 mV; n=4) were not significantly different (p > 0.05). Furthermore, no significant difference was found between the reversal potential of the GTPγS induced outward K+ current (-68 ± 1.1 mV; n=4) and that of IK (-76 ± 3.6 mV; n=6) (p > 0.05).

Molecular biology studies of delayed rectifier type Kv channels showed rapid upregulation of Kv3.1 upon incubation of microglia with Aβ1-42. Aβ1-42 significantly increased relative mRNA of Kv3.1 at 10 min (by 5.6 fold as compared to control p < 0.05; n=3), peaked at 30 min (by 13 fold compared to control p < 0.001) and decreased to non-significant levels at 1 and 2 hrs. All other Kv channels examined were either unchanged (Kv1.1, 1.2, 1.3, 2.1) or decreased (Kv1.5, 1.6). Although the expression of Kv3.1 increased rapidly upon incubation of microglia with Aβ1-42 , this increase in mRNA may not be sufficiently fast to account for the results using patch clamp.

These results indicate that acute Aβ1-42 activates Fc receptors resulting in an immediate transient depolarization of microglia. This depolarization is followed by upregulation of a K+ channel through a G protein and is likely due to Kv3.1 channel activation. The Fc receptor and Kv3.1 channel serve as potential sites for modulation of Aβ1-42 induced microglial inflammatory responses and potential neurotoxicity in the treatment of AD.

Support for this work was provided by a doctoral award to SF from the Alzheimer Society of Canada and research grants to JGM from the Alzheimer Society of Canada and Alzheimer Association USA.