Opioids regulate proliferation of neural progenitors. A study on the effects of opioids on adult rat hippocampal progenitors in vitro and in vivo

Abstract: The opioid receptors were among the first pharmacologically described brain receptors. These opioid receptors have thereafter been shown to mediate effects on proliferation and differentiation in both the embryonal and adult central nervous system. The aim of this thesis was to investigate if cultured hippocampal progenitors from the adult rat brain expressed opioids and opioid receptors and to investigate a possible regulation by opioids on proliferation, differentiation and gene expression in these cultures. Furthermore, we wanted to extend the results and investigate if endogenous opioids regulated hippocampal proliferation in vivo by using opioid receptor antagonists in both non-running and voluntary running rats, a situation associated with increased levels of endogenous opioids. Hippocampal progenitors were found to release b-endorphin that bound to mu- (MOR) and delta- (DOR) but not kappa- (KOR) opioid receptors in vitro. Incubation with MOR or DOR antagonists reduced proliferation whereas stimulation with b-endorphin increased proliferation in these cultures, respectively. The opioid-induced proliferation involved both intracellular calcium and phosphatidylinositol 3-kinase that stimulated phosphorylation of mitogen-activated protein kinase (MAPK). Incubation with naloxone for ten days increased neurogenesis and reduced astrogliogenesis and oligodendrogenesis whereas stimulation with b-endorphin increased oligodendrogenesis but had no effect on astrogliogenesis. Using cDNA arrays, the levels of endogenous opioids were found to regulate gene expression for several cell cycle and oligodendrocyte-specific proteins. Stimulation with b-endorphin also increased Id1, but not Id3 mRNA levels. Down-regulation of Id1 protein using antisense oligonucleotides was suggested to antagonize the opioid-induced oligodendrogenesis.Using bromodeoxyuridine (BrdU), we observed a five-fold increase in hippocampal proliferation after nine days of wheel running in spontaneously hypertensive rats, a rat strain known to run voluntary. This increased BrdU labelling was reduced by peripheral administration of a preferential MOR, but not a DOR antagonist. In non-running rats, MOR and DOR antagonists increased hippocampal proliferation. In non-running, but not in running rats, the opioid receptor antagonists reduced adrenal gland weights and plasma levels of corticosterone. This stress hormone is known to reduce hippocampal neurogenesis. Running rats had three-fold higher levels of hippocampal Met-enkephalin-Arg-Phe levels compared to non-running rats, indicating increased opioid activity in hippocampus during running. These findings demonstrate that the levels of endogenous opioids, acting on MORs and DORs, regulate proliferation of adult rat hippocampal progenitors both in vitro and in vivo. Endogenous opioids were also shown to regulate differentiation and gene expression in cultures of hippocampal progenitors. Finally, we show that exercise robustly increases proliferation of hippocampal progenitors and that such an effect is partly mediated by endogenous opioids.