CNS distribution, signalling properties and central effects of G-protein coupled receptor 4

Abstract

Information on the distribution and biology of the G-protein coupled receptor 4 (GPR4) in the brain is limited. It is currently thought that GPR4 couples to Gs proteins and may mediate central respiratory sensitivity to CO2. Using a knock-in mouse model, abundant GPR4 expression was detected in the cerebrovascular endothelium and neurones of dorsal raphe, retro-trapezoidal nucleus locus coeruleus and lateral septum. A similar distribution was confirmed using RNAscope in situ hybridisation. In HEK293 cells, overexpressing GPR4, it was highly constitutively active at neutral pH with little further increase in cAMP towards acidic pH. The GPR4 antagonist NE 52-QQ57 effectively blocked GPR4-mediated cAMP accumulation (IC50 26.8 nM in HEK293 cells). In HUVEC which natively express GPR4, physiological acidification (pH 7.4–7.0) resulted in a cAMP increase by ∼55% which was completely prevented by 1 μM NE 52-QQ57. The main extracellular organic acid, l-lactic acid (LL; 1–10 mM), suppressed pH dependent activation of GPR4 in HEK293 and HUVEC cells, suggesting allosteric negative modulation. In unanaesthetised mice and rats, NE 52-QQ57 (20 mg kg−1) reduced ventilatory response to 5 and 10% CO2. In anaesthetised rats, systemic administration of NE 52-QQ57 (up to 20 mg kg−1) had no effect on hemodynamics, cerebral blood flow and blood oxygen level dependent responses. Central administration of NE 52-QQ57 (1 mM) in vagotomised anaesthetised rats did not affect CO2-induced respiratory responses. Our results indicate that GPR4 is expressed by multiple neuronal populations and endothelium and that its pH sensitivity is affected by level of expression and LL. NE 52-QQ57 blunts hypercapnic response to CO2 but this effect is absent under anaesthesia, possibly due to the inhibitory effect of LL on GPR4.

Chemosensitivity of Phox2b‐expressing retrotrapezoid neurons is mediated in part by input from 5‐HT neurons

“…However, the role of Gpr4 in chemoreception has recently been called into question (Hosford et al . ). Gpr4 is widely expressed, including in 100% of 5‐HT neurons in the medullary raphe and 83% of C1 adrenergic neurons, as well as by the carotid bodies (Kumar et al .…”

“…Gpr4 is also expressed at high levels in vascular endothelia including in the brain (reviewed by Hosford et al . ), as detected using antibodies, in situ hybridization, GFP expression driven by the Gpr4 promoter, and cre‐mediated GFP expression in cells with a developmental history of Gpr4 expression (Qiao et al . ; Sun et al .…”

Electrochemical carbon fiber-based technique for simultaneous recordings of brain tissue PO2, pH, and extracellular field potentials

“…Briefly, a 72 mm inner diameter volume coil was used for transmission and signal was received using a 4-channel array head coil (Rapid Biomedical). To assess T2* weighted BOLD signals the following sequence parameters were used: TR◻=◻5s, TI◻=◻2s, matrix size◻=◻64◻×◻64, FOV◻=◻35mm × 35mm, TE◻=◻10ms, single slice (slice thickness◻=◻2mm), inversion pulse bandwidth◻=◻20,000Hz (Hosford et al 2018). BOLD responses in the S1FL region were triggered by electrical stimulation of the contralateral forelimb as described above.…”

Analyzing the neuroglial brainstem circuits for respiratory chemosensitivity in freely moving mice

“…The extent to which GPR4 in RTN neurons contributes to respiratory chemosensitivity is the subject of conflicting evidence (30). Global knockout of GPR4 severely attenuates the adaptive response to hypercapnia (8), but this channel is widely expressed centrally and peripherally including in the carotid body chemosensory cells and the medullary raphe (8,30). Our in vivo recordings from freely moving mice suggest that in the rostral chemosensory area, it is the raphe neurons that play the major role in mediating chemosensitivity.…”

“…Recently the pH sensitive channel GPR4 has been claimed to endow RTN neurons with pH sensitivity (8). The extent to which GPR4 in RTN neurons contributes to respiratory chemosensitivity is the subject of conflicting evidence (30). Global knockout of GPR4 severely attenuates the adaptive response to hypercapnia (8), but this channel is widely expressed centrally and peripherally including in the carotid body chemosensory cells and the medullary raphe (8,30).…”