Regular articleReversal of age-related cognitive impairments in mice by an extremely low dose of tetrahydrocannabinol
Introduction
Cognitive decline is an integral aspect of aging. In some cases, cognition deteriorates along the emergence of neurodegenerative maladies such as Alzheimer's, Parkinson's, or Huntington's diseases, while in many other cases it appears as a spontaneous and continuous decline in cognitive competency along normal life. No cure is available for this aggravating condition although preventive measures such as physical exercise, mental activity, and balanced diet are suggested to slow down the process that begins already in midlife. It is widely accepted that at least part of the decline in cognitive functioning reflects the accumulation of minor insults that occur during lifetime, such as short episodes of hypoxia, transient seizures, micro emboli, exposure to neurotoxins, or local inflammatory events within the brain.
In previous studies (Assaf et al., 2011, Fishbein et al., 2012, Fishbein-Kaminietsky et al., 2014), we have shown that ultra-low doses of tetrahydrocannabinol (THC, the main psychotropic ingredient in cannabis) protected young mice from cognitive impairments that were evoked by various insults. Thus, the injection of 0.002 mg/kg of THC (a dose i.e., 3–4 orders of magnitude lower than the doses required to induce the conventional cannabinoid effects in rodents) prevented the long-lasting behavioral deterioration that followed either phenothiazide (“metrazol”)-induced seizures, CO-induced hypoxia, pentobarbital-induced deep anesthesia, methylenedioxymethamphetamine (“ecstasy”)-induced neurotoxicity, or lipopolysaccharide-induced neuroinflammation. A single dose of THC produced a protective effect with a wide therapeutic time window, and the drug could be effectively introduced either 1–7 days before or 1–7 days after the insult. The protective effects of THC that lasted for at least 7 weeks were mediated by CB1 cannabinoid receptors (Fishbein-Kaminietsky et al., 2014) and were accompanied by long-lasting biochemical changes in various regions of the mice brains. For example, the phosphorylation of the kinase extracellular signal-regulated kinase (ERK), the phosphorylation of the transcription factor cyclic adenosine monophosphate response element-binding protein (CREB), and the level of the growth factor brain-derived neurotrophic factor (BDNF) were elevated in the hippocampus and frontal cortex 3–7 weeks following a single injection of THC (Fishbein et al., 2012). It is worth noting that these 3 molecules have been previously shown to be involved in neuroprotection (see e.g., Jin et al., 2006, Lin et al., 2009, Truettner et al., 2002).
In the present study, we tested our hypothesis that a single ultra-low dose of THC can reverse age-dependent cognitive decline in mice. For this purpose, we subjected 24-month-old mice to a battery of behavioral assays that measured different aspects of memory and learning. The experimental data clearly show that a single injection of 0.002 mg/kg of THC eliminated cognitive deficits in old mice and that this beneficial effect lasted for at least 7 weeks. In order to substantiate this surprising behavioral finding, we looked for a possible biochemical substrate within the brain. Sirtuin1 (SIRT1) is an nicotinamide adenine dinucleotide-dependent protein deacetylase that was previously shown to be involved in synaptic plasticity, memory formation, learning capability, neuronal development, and neuroprotection (Gao et al., 2010, Gueguen et al., 2014, Herskovits and Guarente, 2014, Michan et al., 2010, Zhang et al., 2016). SIRT1 was found to mediate the protective effects of resveratrol, melatonin, and calorie restriction (Baur and Sinclair, 2006, Cristofol et al., 2012, Ramis et al., 2015) and was suggested to participate in the pathology of various neurodegenerative diseases (Jiang et al., 2014, Kim et al., 2007). The level of SIRT1 was found to decrease in old mice (Gong et al., 2014, Lafontaine-Lacasse et al., 2010) and in aging humans (Kumar et al., 2013), and its activation reversed cognitive decline in a mouse model of accelerated senescence (Cristofol et al., 2012). In the current study, we found that a single injection of 0.002 mg/kg THC elevated the level of SIRT1-immunoreactive proteins in the hippocampus and in the prefrontal cortex (PFC) of old mice for at least 7 weeks.
We further hypothesized that such long-lasting behavioral and biochemical changes might be accompanied by structural changes in the brain. Magnetic resonance imaging (MRI) is a powerful technique that enables the detection of structural changes in the living brain. Among other parameters, MRI can identify changes in the relative volume of specific brain regions and can indicate changes in the density of the brain tissue. Such changes have previously been detected by MRI in aging subjects (Abe et al., 2002, Head et al., 2004, Pfefferbaum et al., 2005), in neurodegenerative diseases (Bozzali et al., 2002, Song et al., 2004, Yang et al., 2011), and even after short-term behavioral manipulations (Lerch et al., 2011, Scholz et al., 2015). Here, we report that a single application of a low dose of THC induces long-lasting structural alterations in the mouse brain.
Taken together, our behavioral, biochemical, and morphological studies show that extremely low doses of THC can have ameliorating effects and present a feasible treatment to combat age-associated cognitive decline. Preliminary results of this research were previously presented at the 2016 symposium of the International Cannabinoid Research Society (Sarne et al., 2016).
Section snippets
Animals and treatment
All experiments were performed on 70 female Institute of Cancer Research mice: 36 elderly (24 months old) and 12 young (2 months old) mice were used for behavioral experiments; 22 of the old mice were then taken for biochemical examination; another group of 22 old mice that had not been subjected to any behavioral manipulation were used for MRI measurements. The animals were housed 5–10 per cage in the Animal Care Facility at a temperature of 21 °C and a 14/10-hour light/dark cycle, with free
Object and place recognition assays
Three weeks after the injection of either THC (0.002 mg/kg) or vehicle, elderly (24-month-old) mice were introduced to the OR assay. On the habituation day, mice of either group moved freely in the arena. There were no significant differences between the 2 groups in the amount of movement or in the RT they spent in the center of the arena (see Discussion). On day 1 of the OR assay, mice of the 2 groups spent similar time in examining each of the 2 objects. There was no significant difference
Discussion
Our behavioral experiments clearly showed that an acute administration of an ultra-low dose of THC reversed age-associated cognitive impairments in female mice. The effect was not limited to a specific aspect of memory since it was repeated in a variety of assays designed to measure different cognitive functions. Specifically, while the OR, PR, and YM assays depended on the natural tendency of mice to explore and prefer novel stimuli, the Morris' water maze involved learning under stress
Disclosure statement
The authors have no actual or potential conflicts of interest.
Acknowledgements
The authors thank Prof. Raphael Mechoulam of the Hebrew University of Jerusalem for his generous donation of tetrahydrocannabinol. This study was supported by The Mauerberger Foundation, South Africa and by Therapix Biosciences, Israel.
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