Ghrelin promotes reorganization of dendritic spines in cultured rat hippocampal slices

Abstract

Recent evidence suggests ghrelin may up-regulate the number of spine synapses. However, it is not completely understood whether an increased number of synapses are expressed on existing spines or accommodated in newly generated spines. We examined if ghrelin might have promoted the generation of new dendritic spines. Localization of polymerized actin (F-actin), highly expressed in dendritic spines, was assayed using phalloidin, a mushroom toxin that has a high affinity to F-actin. Alexa 488-conjugated phalloidin was visualized and relative changes in fluorescing puncta were quantified using a confocal microscope. Ghrelin was applied to cultured hippocampal slices for either 60 min or 23 h. Ghrelin increased the phalloidin fluorescent signals. The antagonist of the ghrelin receptor, D-Lys3-GHSR-6, blocked the ghrelin’s effect of increasing the phalloidin signal, suggesting that the ghrelin’s effect was mediated by the ghrelin receptor (GHSR1a). The ghrelin-mediated increase in phalloidin signals remained elevated while ghrelin was present in the culture media for 23 h. However, removal of ghrelin from culture media restored the phalloidin signal to control level. Our results suggest ghrelin may have a stimulating effect on the generation or remodeling of dendritic spines, and the spine change persists in the presence of ghrelin. The serum ghrelin level is high when the stomach is empty, and the ghrelin level remains high until metabolic demands are fulfilled. Thus, ghrelin may be involved in food-related and appetite-related learning in the hippocampus.

Ghrelin receptor activity amplifies hippocampal N‐methyl‐d‐aspartate receptor‐mediated postsynaptic currents and increases phosphorylation of the GluN1 subunit at Ser896 and Ser897

“…This novel finding provides a cellular and molecular mechanism to explain earlier reports on the improved memory retention after intra-hippocampal ghrelin application proposed by Carlini et al (2010). It also explains the increased dendritic spine density in hippocampal slice cultures in response to exogenous application of ghrelin (Berrout & Isokawa, 2012), and the enhanced LTP and spine synapse formation induced by systemic application of ghrelin in vivo (Diano et al, 2006). NMDAR function can be regulated by many factors.…”

“…(). It also explains the increased dendritic spine density in hippocampal slice cultures in response to exogenous application of ghrelin (Berrout & Isokawa, ), and the enhanced LTP and spine synapse formation induced by systemic application of ghrelin in vivo (Diano et al ., ).…”

“…The dentate gyrus, CA2 and CA3 show distinctively high expression of GHSR1a mRNA (Guan et al, 1997), and CA1 shows strong binding of biotinylated ghrelin (Diano et al, 2006) and the expression of enhanced green fluorescent protein (EGFP) tagged to GHSR1a (Jiang et al, 2006). We previously reported that exogenous application of ghrelin upregulated the phosphorylation of cAMP response element-binding protein (CREB) (Cuellar & Isokawa, 2011) and enhanced dendritic spine density (Berrout & Isokawa, 2012) through the activation of GHSR1a in the CA1 region of the hippocampus. Diano et al (2006) reported that systemically applied ghrelin crossed the bloodbrain barrier and promoted dendritic spine synapse formation and long-term potentiation in the hippocampus.…”

Ghrelin triggers the synaptic incorporation of AMPA receptors in the hippocampus

“…Previous evidence suggests that GHS-R1a is expressed in the hippocampus (6,(27)(28)(29), but the subcellular localization of GHS-R1a in hippocampal cells has not been studied. Primary hippocampal neurons were immunolabeled at 15 d in vitro (DIV) for endogenous GHS-R1a.…”

Molecular Mechanisms of Drug-Induced Cognitive Enhancement

“…Activation of the ghrelin receptor has also been linked to PKA activation (Kohno et al, 2003), MAPK and PI3K pathways (Camina, 2006), although the mechanisms are still far from clear and may involve heterodimerization with other GPCRs (Schellekens et al, 2013). This effect may be mediated by remodeling of the actin cytoskeleton within dendritic spines (Berrout and Isokawa, 2012;Cuellar and Isokawa, 2011). Thus, it has been shown that circulating ghrelin can enter the hippocampus, where it has a synaptogenic effect in vivo (Diano et al, 2006) and in cortical neuronal cultures (Stoyanova and le Feber, 2014).…”