Growth hormone is protective against acute methadone-induced toxicity by modulating the NMDA receptor complex
Graphical abstract
Introduction
Accumulating evidence indicates that long-term use of opioids such as morphine and methadone can cause cognitive deficits in both animals and humans (Sjogren et al., 2005, Tramullas et al., 2007, Rhodin et al., 2014, Schiltenwolf et al., 2014). These adverse effects are suggested to be associated with neuronal cell death (Mao et al., 2002, Svensson et al., 2008, Perez-Alvarez et al., 2010), reduced neurogenesis (Eisch et al., 2000), and volumetric changes of regions in the brain such as the amygdala, and hypothalamus (Younger et al., 2011). Taken together with the fact that prescription opioid use disorders and frequency of use has increased during 2003–2013 in the US (Han et al., 2015), and that opioid prescription has increased in Scandinavia during 2006–2012 (Mahic et al., 2015), these neurotoxic effects warrant further evaluation.
Recent studies suggest that growth hormone (GH), a somatotrophic hormone released from the anterior pituitary gland, counteracts some of the above-mentioned effects. For instance, GH reduces morphine-induced neurotoxicity in primary hippocampal cell cultures (Svensson et al., 2008), and has been shown to increase overall quality of life and cognitive function in a human patient exposed to chronic methadone treatment (Rhodin et al., 2014). In fact, it is well established that both GH and its mediator insulin-like growth factor-1 (IGF-1) are involved in the protection of the central nervous system (CNS) following injury (see review Nyberg, 2009). The protective effects of GH are demonstrated in various studies, which include GH increasing cell survival after hypoxic-ischemic-induced injury (Scheepens et al., 2001, Alba-Betancourt et al., 2013), preventing apoptosis of lymphocytes (Mitsunaka et al., 2001), and reducing neurological disabilities in mammals (Devesa et al., 2013, Heredia et al., 2013).
In addition to the neuroprotective effects of GH, there is evidence of its role as a cognitive enhancer. First, a relationship between the levels of circulating GH and cognitive function exists, with patients suffering from deficiency of this hormone also having impaired cognitive functions (Falleti et al., 2006). Second, both GH and IGF-1 are strong mediators of both gliogenesis and neurogenesis in the CNS and are thus essential for normal cognitive function (see review Aberg, 2010). Third, GH improves cognitive function in various animal models (Le Greves et al., 2006, Enhamre-Brolin et al., 2013, Gronbladh et al., 2013, Ramis et al., 2013, Studzinski et al., 2015), but also demonstrates promising results in a case study of a human undergoing long-term treatment with methadone (Rhodin et al., 2014).
The relationship between the somatotrophic axis and its beneficial effects in the CNS remains unclear. Nevertheless, evidence indicates that GH increases the transmission of the N-methyl-d-aspartate (NMDA) receptor (see review Nyberg and Hallberg, 2013), a key component for regulating long-term potentiation (LTP) and subsequent facilitation of memory and learning (Tsien et al., 1996). The improved cognitive function following treatment with GH disappears when dizocilpine (MK801), a potent NMDA receptor antagonist, is administered (Ramis et al., 2013). Furthermore, GH increases GluN1, GluN2a, and GluN2b NMDA receptor subunit expression, which has been linked to improved cognitive function (Le Greves et al., 2002, Le Greves et al., 2006, Studzinski et al., 2015). Certain opioids, such as methadone, also have antagonistic affinity to the NMDA receptor (Ebert et al., 1995) marking GH as a promising treatment against methadone-induced injury.
Altogether, evidence suggests that GH is a potent protective agent and acts as a cognitive enhancer in the CNS, but its precise role in these conditions needs to be further evaluated. In this study, we assessed the acute protective effects of recombinant human GH (rhGH) on methadone-induced toxicity in rat primary cortical cell cultures in order to gain further insights into the possible mechanism of rhGH in the brain.
Section snippets
Experimental procedures
All experiments that included animals were approved by the local animal ethics committee at Uppsala University (C14/15) according to the Swedish guidelines regarding animal experiments (Animal Welfare Act SFS1998:56) and the European Communities Council directive (86/609/EEC).
Methadone, MK-801 and morphine, unlike rhGH, induce acute mitochondrial dysfunction
There was an overall effect on mitochondrial function in primary cortical cells for each of the following treatments; methadone, MK801, and morphine (as indicated by ANOVA, p < 0.0001). However, there was no overall effect of rhGH treatment (as indicated by ANOVA, p = 0.1493) shown in Fig. 1A. Post hoc analysis further revealed that methadone significantly reduced the proportion of cells with functional mitochondria at 10–1000 μM in comparison with control (Fig. 1B). The LD50 was calculated to be 60
Discussion
Our main findings from the present study demonstrate that human GH is protective against acute methadone-induced cellular injury. These findings support previous reports indicating that GH acts as a protective agent in the CNS (Scheepens et al., 2001, Svensson et al., 2008, Alba-Betancourt et al., 2013, Devesa et al., 2013, Heredia et al., 2013, Rhodin et al., 2014).
We have examined the protective effects of rhGH using three assays, each measuring specific toxic events. First, the activity of
Conclusion
Overall, we have demonstrated that rhGH may act as a protective agent against methadone-induced cellular injury. Our results further suggest that rhGH may counteract a caspase-independent-mediated cell death (e.g. necrosis), possibly by normalizing or restoring the NMDA receptor complex. In this study, we have provided evidence for a possible mechanism that may contribute to the protective effects of GH. These data encourage further studies, particularly those associated with glial-neuronal
Acknowledgment
This work was supported by the Kjell and Märta Beijer Foundation; the Swedish Research Council (grant number 9459); and the Swedish Brain Foundation. The authors have no conflict of interest to declare.
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