Palmitoylethanolamide Ameliorates Hippocampal Damage and Behavioral Dysfunction After Perinatal Asphyxia in the Immature Rat Brain

Abstract

Perinatal asphyxia (PA) is an obstetric complication associated with an impaired gas exchange. This health problem continues to be a determinant of neonatal mortality and neurodevelopmental disorders. Palmitoylethanolamide (PEA) has exerted neuroprotection in several models of brain injury and neurodegeneration. We aimed at evaluating the potential neuroprotective role of PEA in an experimental model, which induces PA in the immature rat brain. PA was induced by placing Sprague Dawley newborn rats in a water bath at 37°C for 19 min. Once their physiological conditions improved, they were given to surrogate mothers that had delivered normally within the last 24 h. The control group was represented by non-fostered vaginally delivered pups, mimicking the clinical situation. Treatment with PEA (10 mg/kg) was administered within the first hour of life. Modifications in the hippocampus were analyzed with conventional electron microscopy, immunohistochemistry (for NeuN, pNF-H/M, MAP-2, and GFAP) and western blot (for pNF H/M, MAP-2, and GFAP). Behavior was also studied throughout Open Field (OF) Test, Passive Avoidance (PA) Task and Elevated Plus Maze (EPM) Test. After 1 month of the PA insult, we observed neuronal nucleus degeneration in CA1 using electron microscopy. Immunohistochemistry revealed a significant increase in pNF-H/M and decrease in MAP-2 in CA1 reactive area. These changes were also observed when analyzing the level of expression of these markers by western blot. Vertical exploration impairments and anxiety-related behaviors were encountered in the OF and EPM tests. PEA treatment attenuated PA-induced hippocampal damage and its corresponding behavioral alterations. These results contribute to the elucidation of PEA neuroprotective role after PA and the future establishment of therapeutic strategies for the developing brain.

Partial Reversal of Striatal Damage by Palmitoylethanolamide Administration Following Perinatal Asphyxia

“…Fifty-three male rat pups were subjected to PA using an experimental model (Bjelke et al, 1991), which produces moderate to severe PA (Van de Berg et al, 2003;Galeano et al, 2011) as we described previously (Capani et al, 2003(Capani et al, , 2009Saraceno et al, 2010Saraceno et al, , 2012aBlanco et al, 2015;Holubiec et al, 2017;Herrera et al, 2018). The murine model of PA, originally developed by Bjelke et al (1991), has been widely used (Barkhuizen et al, 2017;Herrera et al, 2017b) and accepted to mimic this condition (Capani et al, 1997(Capani et al, , 2003(Capani et al, , 2009Boksa and El-Khodor, 2003;Saraceno et al, 2010Saraceno et al, , 2012aStrackx et al, 2010;Muñiz et al, 2014;Romero et al, 2015;Wakuda et al, 2015;Herrera et al, 2018). The Bjelke model presents several advantages such as follows: (a) asphyxia is produced at birth (Capani et al, 2009); (b) acidosis, hypercapnia, and hypoxia are present in the whole body, replicating a global asphyxia, which is the most common type (Strackx et al, 2010); (c) it is non-invasive, avoiding the effects of surgical procedures; and (d) since hypoxia is produced in the whole body, it affects both brain hemispheres, which makes this model more suitable for behavioral studies (Arteni et al, 2010).…”

“…The Bjelke model presents several advantages such as follows: (a) asphyxia is produced at birth (Capani et al, 2009); (b) acidosis, hypercapnia, and hypoxia are present in the whole body, replicating a global asphyxia, which is the most common type (Strackx et al, 2010); (c) it is non-invasive, avoiding the effects of surgical procedures; and (d) since hypoxia is produced in the whole body, it affects both brain hemispheres, which makes this model more suitable for behavioral studies (Arteni et al, 2010). The control group was represented by nonfostered vaginally delivered pups, mimicking the clinical situation (Herrera et al, 2018).…”

“…PEA (0879/10, Tocris Bioscience, Bristol, United Kingdom) was dissolved in vehicle (VHI), a solution containing 1:1:8 dimethyl sulfoxide, Tween 80, and NaCl and administered by subcutaneous injection in a volume of 10 mg/kg (Herrera et al, 2018). This dose has previously shown to be effective in animal models of experimental spinal cord injury (Genovese et al, 2008), 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-inducedmodel of Parkinson's disease (Esposito et al, 2012), lipopolysaccharideinduced neuroinflammation (Sayd et al, 2014), hippocampal damage and behavioral disturbances induced by PA (Herrera et al, 2018), and cognitive function impairment and astrogliosis induced by neonatal anoxia/ischemia in rats (Holubiec et al, 2018). A preparation similar to that used in this study inhibited the response of mesolimbic dopamine neurons to nicotine in the ventral tegmental area, suggesting the ability of PEA to cross the blood-brain barrier in rats (Melis et al, 2008).…”

“…Compared with other acylethanolamides, PEA showed stronger anti-inflammatory effects reducing PA-induced astrogliosis in P30 rats' hippocampus (Holubiec et al, 2018) and preventing cortical neurons from HI-induced cell death (Portavella et al, 2018). Moreover, PEA treatment during the first hour of life in asphyctic P30 rats reduced hippocampal histological degeneration and prevented the anxiety-like behavior induced by PA (Herrera et al, 2018). However, the protective effects of PEA in the striatum of asphyctic rats have not been addressed.…”

“…Previous studies from our group using an animal model of PA (Bjelke et al, 1991) showed that PA-induced alterations are characterized by increased astrogliosis in the hippocampus and dorsal striatum (Cebral et al, 2006;Saraceno et al, 2010;Holubiec et al, 2017), reduced microtubule-associated protein 2 (MAP-2) level, and alterations in the phosphorylation of high-(200 kDa) and medium-(160 kDa) molecular-weight neurofilaments (pNF-H/M) in the hippocampus in 120-dayold rats (P120, postnatal day 120) (Saraceno et al, 2010) or 30-day-old rats (P30, postnatal day 30) (Herrera et al, 2018). Importantly, MAP-2 is predominantly localized in neuronal dendrites of the caudate, hippocampus, cerebral cortex, Abbreviations: ANOVA, analysis of variance; CNS, central nervous system; CO, carbon monoxide; GFAP, glial fibrillary acidic protein; HI, hypoxia-ischemia; HIE, hypoxia-ischemia encephalopathy; MAP-2, microtubule-associated protein-2; MPTP, 1-methyl-4-phenyl-1,2,3,6tetrahydropyridine; PEA, palmitoylethanolamide; PA, perinatal asphyxia; PPARα, peroxisome proliferator-activated receptor alpha; pNF-H/M, phosphorylated high/medium-molecular-weight NF.…”