Reduced Hyperpolarization-Activated Current Contributes to Enhanced Intrinsic Excitability in Cultured Hippocampal Neurons from PrP(-/-) Mice.

Jing Fan,Maria A Gandini,Zizhen Zhang,Ivana A Souza,Patrick L Stemkowski,Lina Chen,Stefanie A Black,Gerald W Zamponi

doi:10.3389/fncel.2016.00074

Abstract

Genetic ablation of cellular prion protein (PrPC) has been linked to increased neuronal excitability and synaptic activity in the hippocampus. We have previously shown that synaptic activity in hippocampi of PrP-null mice is increased due to enhanced N-methyl-D-aspartate receptor (NMDAR) function. Here, we focused on the effect of PRNP gene knock-out (KO) on intrinsic neuronal excitability, and in particular, the underlying ionic mechanism in hippocampal neurons cultured from P0 mouse pups. We found that the absence of PrPC profoundly affected the firing properties of cultured hippocampal neurons in the presence of synaptic blockers. The membrane impedance was greater in PrP-null neurons, and this difference was abolished by the hyperpolarization-activated cyclic nucleotide-gated (HCN) channel blocker ZD7288 (100 μM). HCN channel activity appeared to be functionally regulated by PrPC. The amplitude of voltage sag, a characteristic of activating HCN channel current (Ih), was decreased in null mice. Moreover, Ih peak current was reduced, along with a hyperpolarizing shift in activation gating and slower kinetics. However, neither HCN1 nor HCN2 formed a biochemical complex with PrPC. These results suggest that the absence of PrP downregulates the activity of HCN channels through activation of a cell signaling pathway rather than through direct interactions. This in turn contributes to an increase in membrane impedance to potentiate neuronal excitability.

Highlights

Cellular prion protein (PrPC) is a naturally occurring protein, whose abnormal conformation can lead to a range of neurological disorders including Creutzfeldt-Jakob disease, Kuru and bovine spongiform encephalitis (Knight and Will, 2004)
We used intracellular recordings from cultured hippocampal pyramidal neurons to characterize the effect of PrPC on hippocampal neuron intrinsic excitability in the presence of synaptic blockers DNQX (20 μM) and D,L-APV (50 μM)
We found that the absence of PrPC strongly affected firing properties of hippocampal neurons, increasing number of action potential (AP) (Figures 1A,B) and decreasing the spike threshold (Figure 1C) in response to 250 ms depolarizing step current injections (369.2 ± 28.6 pA for WT (n = 13) vs. 140.0 ± 14.5 pA for KO (n = 10), p < 0.001)

Summary

Introduction

Cellular prion protein (PrPC) is a naturally occurring protein, whose abnormal conformation can lead to a range of neurological disorders including Creutzfeldt-Jakob disease, Kuru and bovine spongiform encephalitis (Knight and Will, 2004) This abnormal protein accumulates in the brain, forming polymeric aggregates that disrupt synapses, leading to loss of dendrites and neuronal death (Soto and Satani, 2011). Recordings from PrP-null neurons in vitro showed a variety of electrophysiological abnormalities, with reduced Ca2+dependent K+ currents (Colling et al, 1996), abnormal type-A γ-aminobutyric acid receptor (GABAAR) activity (Collinge et al, 1994), as well as a significant reduction of afterhyperpolarization potentials (AHP; Colling et al, 1996; Herms et al, 2001; Mallucci et al, 2002; Fuhrmann et al, 2006) Despite this progress, we do not yet fully understand how PrPC regulates neuronal output

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