Different patterns of Ca²⁺ signals are induced by low compared to high concentrations of P2Y agonists in microglia

Area of Science:

• Neuroscience
• Immunology
• Cell Biology

Background:

• Microglia, the brain's resident macrophages, play dual roles in maintaining tissue integrity and mediating inflammatory responses.
• Nucleotides are increasingly recognized as critical regulators of microglial functions.
• P2Y receptors are implicated in nucleotide signaling pathways within microglia.

Purpose of the Study:

• To define the expression profile of P2Y receptors in rat microglia.
• To characterize the signaling mechanisms of P2Y receptors, focusing on calcium (Ca²⁺) mobilization.
• To investigate the differential effects of nucleotide concentration and type on microglial Ca²⁺ responses.

Main Methods:

• Utilized rat microglia for experimental studies.
• Applied endogenous nucleotides at various concentrations (100 µM, sub-micromolar) to stimulate P2Y receptors.
• Employed pharmacological agents and pertussis toxin to investigate signaling pathways, including G protein involvement (Gi, Gq).
• Monitored Ca²⁺ transients using fluorescence-based assays.

Main Results:

• Endogenous nucleotides at 100 µM induced robust Ca²⁺ transients via P2Y₂, P2Y₆, and P2Y₁₂ receptors.
• Adenosine diphosphate (ADP) signaling through P2Y₁₂ receptors was confirmed via pharmacological sensitivity and pertussis toxin treatment.
• Adenine and uracil nucleotides differentially activated G proteins (Gi and Gq, respectively) and induced distinct Ca²⁺ responses, including capacitative Ca²⁺ plateau by uracil nucleotides.
• Low concentrations of uracil nucleotides elicited low-frequency oscillatory Ca²⁺ changes, sometimes with sustained extracellular Ca²⁺-dependent rises.

Conclusions:

• Different patterns of Ca²⁺ transients are induced by varying nucleotide concentrations, suggesting distinct functional outcomes.
• The type and concentration of nucleotides modulate microglial responses through specific P2Y receptor signaling pathways.
• These findings highlight the complex role of nucleotide signaling in regulating microglial functions, with implications for brain homeostasis and pathology.