Minocycline prevents neuronal hyperexcitability and neuroinflammation in medial prefrontal cortex, as well as memory impairment caused by repeated toluene inhalation in adolescent rats
Introduction
Inhalant misuse is the intentional inhalation of high concentrations (several thousand ppm) of volatile compounds to achieve psychoactive effects (Balster et al., 2009; Lubman et al., 2008). This practice is particularly prevalent among adolescents and young adults (Crossin et al., 2019; Hynes-Dowell et al., 2011; Villatoro et al., 2011). Toluene is the most commonly and better characterized misused inhalant in terms of behavioral, molecular and electrophysiological effects (Cruz et al., 2014; Shelton, 2016; Win-Shwe and Fujimaki, 2010; Woodward and Beckley, 2014). Preclinical studies have shown that acute exposure to toluene, at concentrations relevant to solvent misuse, produces a wide range of behavioral responses, most of which are typical of central nervous system depressant drugs (Cruz et al., 2014), including memory impairment (Dick et al., 2014; Huerta-Rivas et al., 2012). As for its mechanism of action, toluene releases dopamine (Beckley and Woodward, 2013), acts as a positive allosteric modulator of GABAA, glycine and 5HT3 receptors (Beckstead et al., 2000; Lopreato et al., 2003), is a non-competitive, highly selective NMDA receptor antagonist (Bale et al., 2005; Cruz et al., 1998), and a sodium (Cruz et al., 2003), calcium (Tillar et al., 2002) and potassium channel blocker (Del Re et al., 2006), among other effects (Beckley and Woodward, 2013; Bowen et al., 2006; Lo et al., 2009).
We have recently shown that repeated exposure to 8000 ppm (30 min, 2×/day, for 10 days), but not to lower concentrations, increases the excitability of medial prefrontal cortex (mPFC) pyramidal neurons (Armenta-Resendiz et al., 2018). Similar findings were reported by Wayman and Woodward after a brief exposure to 10,500 ppm toluene (Wayman and Woodward, 2017). The mPFC is involved in decision-making, working memory, attention, planning, cognitive flexibility, learning and impulse control (Euston et al., 2012). This brain area is particularly vulnerable during adolescence to different insults, including drugs of abuse (Goldstein and Volkow, 2002; Winters and Arria, 2011).
Toluene is metabolized to hippuric acid mainly via cytochrome P450 2E1 (CYP2E1) and produces reactive oxygen species (ROS) through various mechanisms (Kodavanti et al., 2015; Tamie and Rui-Sheng, 1994). ROS can be damaging to protein structure and DNA material. Moreover, oxidative stress leads to a reactive state of microglia and astrocytes, as well as the release of pro-inflammatory mediators (Qin and Crews, 2012), including interleukin-1β (IL-1β) (Gustafson-Vickers et al., 2008; He et al., 2019). IL-1β synthesis depends on the activation of transcription factor NF-κB (nuclear factor enhancer of the kappa light chains of activated B cells). ROS overproduction modulates NF-κB, while ROS inhibition prevents inflammasome expression (Abais et al., 2015; Juliana et al., 2012; Morgan and Liu, 2011). Inflammasomes are cytosolic multiprotein complexes with a sensor protein of the NOD-like receptor (NLR) family, needed for inflammatory cytokines' activation and caspase-1 processing (Guo et al., 2015). In particular, activation of NLRP3 (NLR containing pyrin domain 3) is needed for maturation of pro-IL-1β into IL-1β (Cullen et al., 2015; Martin, 2016). In experiments simulating toluene as an occupational hazard, long-exposure periods to 1500 ppm for 4 h per day for 7 days (Gotohda et al., 2002) or 0–500 ppm toluene for 6 weeks (Win-Shwe et al., 2011) produce an increase in pro-inflammatory cytokines and NF-κB mRNA in rodent hippocampus and cerebellum. In addition, Kanter and colleagues found evidence of dilated cisternae of the endoplasmic reticulum and swelling of intracellular mitochondria (which may signify neuroinflammation) in rats exposed to 3000 ppm toluene, 8 h per day, 6 days per week for 12 weeks (Kanter, 2008). To our knowledge, there are no studies addressing the possibility that binge patterns of repeated toluene exposure, mimicking solvent misuse, can also produce neuroinflammatory effects.
Minocycline (MN) is a potent anti-inflammatory agent (Blum et al., 2004; Choi et al., 2005; Dean et al., 2012; Ray et al., 2014) that inhibits the NF-κB pathway and confers neuroprotection by reducing cytokine and ROS production (Blum et al., 2004; Henry et al., 2008; Tikka and Koistinaho, 2001). By utilizing this pharmacological agent, we can indirectly assess oxidative stress produced by repeated toluene exposure. Based on these findings, the aim of this work was two-fold: i) to evaluate if neuroinflammation occurs in the mPFC of adolescent rats repeatedly exposed to a high toluene concentration; and ii) to test the hypothesis that MN would prevent the effects of repeated toluene exposure on neuronal hyperexcitability, neuroinflammation and memory impairment.
Section snippets
Animals
We used a total of 124 adolescent male Wistar rats (25 days of age at the beginning of the experiment) from our own vivarium. Animals were socially housed, had ad libitum access to food and water, and were kept under a 12:12 h light/dark cycle in a climate-controlled room at 22 ± 2 °C. Experimental procedures were carried out in strict accordance with the Mexican Official Norm for utilization and care of laboratory animals “NOM-062-ZOO-1999” and the Institutional Ethics Committee (Protocols
Effect of minocycline on the intrinsic excitability and firing frequency of mPFC layer V prelimbic neurons
Using whole cell recordings, we first aimed to establish whether the daily administration of MN modified the neurophysiological properties of prelimbic mPFC neurons and found that there were no significant differences between the resting membrane potential (RMP) of control and MN-treated neurons (RMP in control = −72.5 ± 1.3 mV; MN group = −70.8 ± 1.9 mV; P = .49, Student's t-test). Then, we constructed a series of I-V relationships. The trajectory and rectification properties of the I-V plots
Minocycline does not significantly alter basal properties of mPFC neurons
Consistent with previous reports (Armenta-Resendiz et al., 2018; Cruz et al., 2019), we showed that repeated exposure to toluene, at 8000 ppm, a concentration relevant to solvent intoxication, increases the firing output of mPFC neurons in the adolescent rat brain. This effect was prevented when animals exposed to toluene were consecutively treated with MN. This drug is an antibiotic and a very effective anti-inflammatory agent (Garrido-Mesa et al., 2013; Möller et al., 2016). Because of its
Conclusion
A binge pattern of toluene exposure produces neuroinflammation and hyperexcitability of layer 5 neurons of prelimbic mPFC, as well as memory impairment in two assays related to PFC function more generally. The finding that the anti-inflammatory antibiotic MN can prevent these effects suggests that neuroinflammation causes several chronic toluene effects most probably via ROS generation and NF-κB pathway activation. These results highlight the risks of repeated exposure to high toluene
Disclosure statement
Silvia L. Cruz reports grant 239192 from Conacyt (National Council of Science and Technology), Mexico. Monserrat Armenta-Reséndiz and César J. Carranza-Aguilar report scholarships from Conacyt, Mexico. Emilio J. Galván has nothing to disclose.
Declaration of Competing Interest
Funding was provided by grant 239192 (SLC); CB 2016-281617 (EJG) and scholarships 261795 (MAR) and 338376 (CJCA) from Conacyt (National Council of Science and Technology) Mexico.
Acknowledgements
This paper includes data from MAR's Doctoral Dissertation. We thank Araceli Hernández-Mendoza for technical support and Eduardo Violante for some artwork design.
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Systemic administration of lipopolysaccharide induces hyperexcitability of prelimbic neurons via modulation of sodium and potassium currents
2022, NeuroToxicologyCitation Excerpt :Consistent with this tenet, previous studies have documented the effects of de novo synthesized proinflammatory cytokines on the function of voltage-gated ion channels that control neuronal excitability and synaptic plasticity (for review, see Viviani et al., 2007; Schäfers and Sorkin, 2008; Vezzani and Viviani, 2015). For instance, a recent study in the rat prefrontal cortex demonstrated that repeated exposure to toluene increases the expression of the proinflammatory interleukin IL-1β, resulting in increased intrinsic excitability of RS PL PC (Cruz et al., 2020). Other studies have characterized the biochemical and biophysical alterations of multiple ion channels in the prefrontal cortex induced by maternal immune activation, an inflammation-mediated model of schizophrenia (Mi et al., 2019).
A novel preclinical model of environment-like combined benzene, toluene, ethylbenzene, and xylenes (BTEX) exposure: Behavioral and neurochemical findings
2022, Neurotoxicology and TeratologyCitation Excerpt :The significant effects reported by Wang et alia in their model of indoor FBTX exposure versus our BTEX model may be due to the greater neurotoxicity of formaldehyde over ethylbenzene (ATSDR, 2010; Pitten et al., 2000), longer exposures (90 vs 15 days), measuring whole-brain monoamines rather than mPFC and NAc alone, and/or greater difficulty of the Morris Maze over the Y-maze. The minimal effects of repeated BTEX exposure in the present study are also in contrast to previous preclinical reports reporting significant behavioral changes in effects on anxiety-like behaviors, like defensive burying and passive avoidance, after acute exposures after individual benzene, toluene, ethylbenzene, or xylene at either abuse-like patterns (higher-concentrations, shorter exposures) (Armenta-Resendiz et al., 2019; Cruz et al., 2020; Molnar et al., 1986; Tegeris and Balster, 1994) or after more environmental-like paradigms (lower-concentrations, much longer exposures) (Berenguer et al., 2004; Berenguer et al., 2003; Korsak et al., 1992; Niaz et al., 2015; von Euler et al., 1991; von Euler et al., 1994; von Euler et al., 2000; Wang et al., 2014; Win-Shwe et al., 2012). While our not restricting access to food before Y-maze testing may have avoided interactions with BTEX exposure, it is also possible that manipulating motivation could have revealed differences between exposed and non-exposed groups.
- 1
SLC and EJG contributed equally to this work.
- 2
Present address: Department of Neuroscience, Medical University of South Carolina, SC, USA.