Epigenetic changes coincide with in vitro primate GnRH neuronal maturation

Area of Science:

• Neuroscience
• Developmental Biology
• Epigenetics

Background:

• Pulsatile gonadotropin-releasing hormone (GnRH) release is crucial for reproduction but its underlying cellular and molecular mechanisms are poorly understood.
• Investigating GnRH neuron development offers insights into neuroendocrine regulation and potential causes of reproductive disorders.

Purpose of the Study:

• To examine developmental changes in intracellular calcium dynamics, peptide release, gene expression, and DNA methylation in cultured rhesus monkey GnRH neurons.
• To elucidate the molecular mechanisms, particularly epigenetic modifications, that regulate GnRH neuron maturation and function.

Main Methods:

• Cultured GnRH neurons from rhesus monkey nasal placodes were studied over 20 days in vitro (div).
• Intracellular calcium ([Ca(2+)](i)) dynamics, GnRH peptide release, GnRH mRNA levels, and GnRH gene CpG methylation status were measured.
• Bisulfite sequencing was used to analyze DNA methylation patterns across a 5' CpG island of the GnRH gene.

Main Results:

• Immature GnRH neurons (≤12 div) showed limited calcium fluctuations and sparse GnRH release.
• Mature GnRH neurons (14-18 div) exhibited synchronized [Ca(2+)](i) oscillations and GnRH pulses at ~60-min intervals.
• GnRH mRNA levels increased significantly (P < 0.05) between 14-20 div, coinciding with maximal peptide release and significant DNA demethylation (P < 0.05) at specific CpG sites.

Conclusions:

• Epigenetic differentiation, specifically DNA demethylation, occurs during GnRH neuronal development in rhesus monkeys.
• Increased GnRH gene expression and decreased CpG methylation are molecular phenotypes associated with mature GnRH neurons.
• This study provides the first evidence of developmental DNA demethylation in postmitotic neurons contributing to a stable neuronal phenotype.