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VIP receptor subtypes in mouse cerebral cortex: evidence for a differential localization in astrocytes, microvessels
J L Martin1, D L Feinstein, N Yu
1Institut de Physiologie, Faculté de Médecine, Université de Lausanne, Switzerland.
Abstract:
The binding characteristics of a monoiodinated form of vasoactive intestinal peptide (M-[125I]VIP) to the membranes of astrocytes, intraparenchymal microvessels and synaptosomes were analyzed in mouse cerebral cortex. Binding to astrocytes, studied in primary cultures, indicates the presence of a single class of high affinity binding sites with a Kd of 3.3 nM and a Bmax of 565 fmol/mg protein. The structurally related peptide secretin does not compete for sites labeled by M-[125I]VIP. In cultured astrocytes, VIP has been previously shown to promote glycogenolysis. Secretin, despite its lack of interaction with sites labeled by M-[125I]VIP, stimulates glycogenolysis with an EC50 of 0.5 nM, thus demonstrating the presence in astrocytes of functional secretin receptors independent from those for VIP. Trypsinization of the primary astrocyte cultures followed by replating as secondary cultures, reveals a second class of low affinity binding sites, with a Kd of 41.3 nM and a Bmax of 881 fmol/mg protein. Secretin does not compete for this class of low affinity binding sites either. Binding of M-[125I]VIP to intraparenchymal microvessels reveals the presence of two classes of binding sites with Kd of 1.4 and 30.3 nM, and Bmax of 7.1 and 73.8 pmol/mg protein, respectively. Similar to what is observed in primary or secondary astrocyte cultures, secretin does not interact with these sites. In this cell type VIP stimulates cAMP formation with an EC50 of 18 nM, while secretin is ineffective. Finally, in agreement with previous reports in rat and guinea pig cerebral cortex, two classes of binding sites are observed in synaptosomal membranes: a high affinity class with a Kd of 4.9 nM and a Bmax of 316 fmol/mg protein, and a low affinity class with a Kd of 42.8 nM and a Bmax of 1578 fmol/mg protein. In contrast to what is observed in non-neuronal membranes, in synaptosomal membranes, secretin effectively competes for sites labeled by M-[125I]VIP with an EC50 of approximately 150 nM. These results indicate that secretin may represent a useful tool to discriminate between neuronal and non-neuronal VIP binding sites, since it competes with M-[125I]VIP exclusively for the neuronal class of binding sites.
Insights
This study analyzed vasoactive intestinal peptide (VIP) binding sites in mouse brain cells. Secretin, a related peptide, distinguishes neuronal VIP sites from non-neuronal ones, aiding research in brain function and disease.
Area of Science:
- Neuroscience
- Molecular Biology
- Biochemistry
Background:
- Vasoactive intestinal peptide (VIP) is a neuropeptide with diverse physiological roles.
- Understanding VIP receptor distribution is crucial for elucidating its functions in the central nervous system.
- Differentiating between neuronal and non-neuronal VIP binding sites can clarify specific cellular responses.
Purpose of the Study:
- To characterize the binding properties of vasoactive intestinal peptide (VIP) in different mouse cerebral cortex cell types.
- To investigate the potential of secretin as a tool to differentiate between neuronal and non-neuronal VIP binding sites.
- To analyze VIP binding site characteristics in astrocytes, microvessels, and synaptosomes.
Main Methods:
- Radioligand binding assays using monoiodinated VIP (M-[125I]VIP).
- Primary and secondary astrocyte cultures were used to study non-neuronal binding.
- Analysis of binding in intraparenchymal microvessels and synaptosomal membranes.
- Competition binding studies with secretin to assess receptor specificity.
Main Results:
- Astrocytes exhibit a single high-affinity class of VIP binding sites, with secretin showing no competition.
- Intraparenchymal microvessels display two classes of VIP binding sites, also unresponsive to secretin.
- Synaptosomes possess two classes of VIP binding sites, where secretin competes for M-[125I]VIP binding.
- Secretin effectively competes for neuronal VIP binding sites but not for those on astrocytes or microvessels.
Conclusions:
- Secretin serves as a valuable pharmacological tool to distinguish neuronal VIP binding sites from non-neuronal ones in the cerebral cortex.
- The distinct binding profiles of VIP and secretin in different cell types highlight receptor heterogeneity.
- These findings contribute to a better understanding of VIPergic neurotransmission and potential therapeutic targets.