I_h blockade reduces cocaine-induced firing patterns of putative dopaminergic neurons of the ventral tegmental area in the anesthetized rat

Highlights

• I_h blockade diminishes VTA DA neuronal spontaneous firing activity.

• I_h inhibition after acute cocaine administration increases the interspike interval.

• I_h reduction can oppose cocaine enhanced VTA DA cell's excitability.

Abstract

The hyperpolarization-activated cation current (I_h) is a determinant of intrinsic excitability in various cells, including dopaminergic neurons (DA) of the ventral tegmental area (VTA). In contrast to other cellular conductances, I_h is activated by hyperpolarization negative to -55 mV and activating I_h produces a time-dependent depolarizing current. Our laboratory demonstrated that cocaine sensitization, a chronic cocaine behavioral model, significantly reduces I_h amplitude in VTA DA neurons. Despite this reduction in I_h, the spontaneous firing of VTA DA cells after cocaine sensitization remained similar to control groups. Although the role of I_h in controlling VTA DA excitability is still poorly understood, our hypothesis is that I_h reduction could play a role of a homeostatic controller compensating for cocaine-induced change in excitability. Using in vivo single-unit extracellular electrophysiology in isoflurane anesthetized rats, we explored the contribution of I_h on spontaneous firing patterns of VTA DA neurons. A key feature of spontaneous excitability is bursting activity; bursting is defined as trains of two or more spikes occurring within a short interval and followed by a prolonged period of inactivity. Burst activity increases the reliability of information transfer. To elucidate the contribution of I_h to spontaneous firing patterns of VTA DA neurons, we locally infused an I_h blocker (ZD 7288, 8.3 μM) and evaluated its effect. I_h blockade significantly reduced firing rate, bursting frequency, and percent of spikes within a burst. In addition, I_h blockade significantly reduced acute cocaine-induced spontaneous firing rate, bursting frequency, and percent of spikes within a burst. Using whole-cell patch-clamp, we determine the progressive reduction of I_h after acute and chronic cocaine administration (15 mg/k.g intraperitoneally). Our data show a significant reduction (~25%) in I_h amplitude after 24 but not 2 h of acute cocaine administration. These results suggest that a progressive reduction of I_h could serve as a homeostatic regulator of cocaine-induced spontaneous firing patterns related to VTA DA excitability.

Introduction

Hyperpolarization-activated cation current (I_h) is a significant modulator of intrinsic excitability on neurons of the mesocorticolimbic system, including dopaminergic cells (DA) from the ventral tegmental area (VTA) (DiFrancesco, 1981; Adams and Halliwell, 1982; Kase and Imoto, 2012). Neuroadaptations in this network are hypothesized to trigger substance abuse disorder. In contrast to other ionic conductances, I_h is a slowly activating cation inward current activated by hyperpolarizing inputs negative to -55 mV. I_h activation depolarizes the membrane to a threshold level for the generation of action potentials (Tsantoulas et al., 2016). Additionally, I_h is involved in various neuronal properties such as the control and modulation of the rhythmic activity of neural circuits (Leresche et al., 1990; McCormick and Pape, 1990), regulation of the resting membrane potential (Doan and Kunze, 1999), firing frequency modulation (Friedman, 2014; Okamoto et al., 2006), regulation of synaptic transmission (Beaumont and Zucker, 2000; Sparks and Chapman, 2014) and dendritic integration of synaptic inputs (Arencibia-Albite et al., 2017; Engel and Seutin, 2015; Magee, 2000). On VTA DA neurons, I_h is classified as an excitatory driving force (Santoro and Shah, 2020; Zhong et al., 2017; Wanat et al., 2008; Neuhoff et al., 2002).

Neuromodulation of intrinsic properties is a decisive determinant of neuron excitability (Desai et al., 1999; Nelson et al., 2003). Intrinsic excitability is a measure of the neuron's inherent electrical properties that can be adjusted by numerous factors (Kemenes et al., 2006; O'Leary, 2010). These adjustments arise through changes in morphological features and the expression level or biophysical properties of ion channels in the membrane (Williams et al., 2013). The activity of a single neuron results from the summation of excitatory and inhibitory synaptic inputs, and the intrinsic membrane properties (Beck and Yaari, 2008). Neuroadaptations can be categorized as either homeostatic, that contribute to stabilizing neuronal excitability, or non-homeostatic, that produce an alteration from basal excitability (Desai et al., 1999; Howard et al., 2007; Wijesinghe and Camp, 2011). Homeostatic changes in neuronal excitability can arise in response to long periods of altered activity (Turrigiano Gina, 1999; Turrigiano and Nelson, 2000; Keck, 2017). For example, 3 h after acute cocaine administration, activity of VTA DA neurons recorded in vivo, is significantly increased (Creed et al., 2016). Our laboratory demonstrated that cocaine sensitization, a chronic cocaine behavioral model, significantly reduces I_h amplitude by ~40% in VTA DA neurons (Arencibia-Albite et al., 2012). This reduction of current occurs concomitantly with the reduction of membrane surface HCN2 protein, the main HCN subunit in VTA DA neurons (Santos-Vera, 2019). An important observation is that after the development of cocaine sensitization, the spontaneous firing of VTA DA cells remains similar to that of control groups (Arencibia-Albite et al., 2012). These findings suggest that I_h reduction could reduce cocaine-induced excitability as a homeostatic adaptation to regulate neuronal excitability of VTA DA neurons.

I_h increases burst firing by enhancing rebound spiking after the disinhibition of VTA DA neurons (Tateno and Robinson, 2011). Additionally, pharmacological blockade of this current on VTA DA neurons can significantly decrease (by 40%) the firing rate in the majority of these neurons; this reduction is suggested to be behaviorally significant (Seutin et al., 2001). While this study indicates that the modulation of I_h can have a role in controlling VTA DA neuronal excitability, the relationship is still poorly understood. Hence, the study of I_h modulation on spontaneous firing activity could lead us to elucidate its contribution to VTA DA neuronal excitability.

To determine if reduction of I_h can alter basal VTA DA neuronal excitability, we evaluated the effect of I_h blockade on spontaneous firing patterns using in vivo single-unit extracellular recordings. VTA DA neurons can be divided by their firing patterns, classified by the average firing rate and the percentage of spike within a burst (%SWB) (Mameli-Engvall et al., 2006). To investigate how I_h modulates cocaine-dependent excitability, we measured I_h using whole-cell patch-clamp electrophysiology, 2 and 24 h after acute cocaine injection. Additionally, we determined the effect of I_h blockade on cocaine-induced spontaneous firing patterns using in vivo single-unit extracellular recordings. We postulate that a progressive I_h reduction serves as a homeostatic regulator to oppose cocaine-induced excitability in VTA DA neurons.