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Distribution and characterisation of Glucagon-like peptide-1 receptor expressing cells in the mouse brain.

Cork SC, Richards JE, Holt MK, Gribble FM, Reimann F, Trapp S - Mol Metab (2015)

Bottom Line: Large numbers of eYFP or tdRFP immunoreactive cells were found in the circumventricular organs, amygdala, hypothalamic nuclei and the ventrolateral medulla.However, tdRFP positive neurons were also found in areas without preproglucagon-neuronal projections like hippocampus and cortex.GLP-1R expression was confirmed in whole-cell recordings from BNST, hippocampus and PVN, where 100 nM GLP-1 elicited a reversible inward current or depolarisation.

View Article: PubMed Central - PubMed

Affiliation: Centre for Cardiovascular and Metabolic Neuroscience, Department of Neuroscience, Physiology & Pharmacology, University College London, London, WC1E 6BT, UK.

ABSTRACT

Objective: Although Glucagon-like peptide 1 is a key regulator of energy metabolism and food intake, the precise location of GLP-1 receptors and the physiological relevance of certain populations is debatable. This study investigated the novel GLP-1R-Cre mouse as a functional tool to address this question.

Methods: Mice expressing Cre-recombinase under the Glp1r promoter were crossed with either a ROSA26 eYFP or tdRFP reporter strain to identify GLP-1R expressing cells. Patch-clamp recordings were performed on tdRFP-positive neurons in acute coronal brain slices from adult mice and selective targeting of GLP-1R cells in vivo was achieved using viral gene delivery.

Results: Large numbers of eYFP or tdRFP immunoreactive cells were found in the circumventricular organs, amygdala, hypothalamic nuclei and the ventrolateral medulla. Smaller numbers were observed in the nucleus of the solitary tract and the thalamic paraventricular nucleus. However, tdRFP positive neurons were also found in areas without preproglucagon-neuronal projections like hippocampus and cortex. GLP-1R cells were not immunoreactive for GFAP or parvalbumin although some were catecholaminergic. GLP-1R expression was confirmed in whole-cell recordings from BNST, hippocampus and PVN, where 100 nM GLP-1 elicited a reversible inward current or depolarisation. Additionally, a unilateral stereotaxic injection of a cre-dependent AAV into the PVN demonstrated that tdRFP-positive cells express cre-recombinase facilitating virally-mediated eYFP expression.

Conclusions: This study is a comprehensive description and phenotypic analysis of GLP-1R expression in the mouse CNS. We demonstrate the power of combining the GLP-1R-CRE mouse with a virus to generate a selective molecular handle enabling future in vivo investigation as to their physiological importance.

No MeSH data available.


Related in: MedlinePlus

GLP-1 elicits electrical responses in GLP-1R-Cre positive cells in various brain regions. A, B. Voltage-clamp recording from an RFP-positive PVN neuron. Bath application of 100 nM GLP-1 elicited a small inward current at a holding potential of −50 mV accompanied by an increase in whole-cell conductance. These effects reversed upon washout of GLP-1 from the bath solution. C, D. Similarly, in a BNST neuron bath application of 100 nM GLP-1 elicits an inward current accompanied by an increase in conductance. E. Voltage clamp recordings in RFP-positive hippocampal neurons revealed that GLP-1 either elicits an inward current (left panel) as in PVN and BNST, or an outward current. F. Mean data from current clamp recordings (left panel) demonstrating that GLP-1 causes a 10–15 mV depolarisation in most neurons tested, and a hyperpolarisation in individual hippocampal cells. Mean data from voltage clamp recordings in the right panel demonstrate the inward current of 10–20 pA amplitude elicited by 100 nM GLP-1 underlying the depolarisation in current clamp. n-numbers are given below the bars. **: p < 0.01.
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fig7: GLP-1 elicits electrical responses in GLP-1R-Cre positive cells in various brain regions. A, B. Voltage-clamp recording from an RFP-positive PVN neuron. Bath application of 100 nM GLP-1 elicited a small inward current at a holding potential of −50 mV accompanied by an increase in whole-cell conductance. These effects reversed upon washout of GLP-1 from the bath solution. C, D. Similarly, in a BNST neuron bath application of 100 nM GLP-1 elicits an inward current accompanied by an increase in conductance. E. Voltage clamp recordings in RFP-positive hippocampal neurons revealed that GLP-1 either elicits an inward current (left panel) as in PVN and BNST, or an outward current. F. Mean data from current clamp recordings (left panel) demonstrating that GLP-1 causes a 10–15 mV depolarisation in most neurons tested, and a hyperpolarisation in individual hippocampal cells. Mean data from voltage clamp recordings in the right panel demonstrate the inward current of 10–20 pA amplitude elicited by 100 nM GLP-1 underlying the depolarisation in current clamp. n-numbers are given below the bars. **: p < 0.01.

Mentions: Seven red-fluorescent PVN neurons were recorded in voltage clamp at a holding potential of −50 mV, and five cells were recorded in current-clamp mode. The mean resting potential in current-clamp mode was −57 ± 2 mV (n = 5). Bath-application of 100 nM GLP-1 caused a depolarisation by 14 ± 1 mV in all five cells that reversed upon washout of the drug. In voltage clamp GLP-1 application induced a reversible inward current of 16 ± 3 pA (n = 7) that was accompanied by a decrease in membrane resistance from 1.1 ± 0.2 GΩ to 0.8 ± 0.1 GΩ (Figure 7A–C, F).


Distribution and characterisation of Glucagon-like peptide-1 receptor expressing cells in the mouse brain.

Cork SC, Richards JE, Holt MK, Gribble FM, Reimann F, Trapp S - Mol Metab (2015)

GLP-1 elicits electrical responses in GLP-1R-Cre positive cells in various brain regions. A, B. Voltage-clamp recording from an RFP-positive PVN neuron. Bath application of 100 nM GLP-1 elicited a small inward current at a holding potential of −50 mV accompanied by an increase in whole-cell conductance. These effects reversed upon washout of GLP-1 from the bath solution. C, D. Similarly, in a BNST neuron bath application of 100 nM GLP-1 elicits an inward current accompanied by an increase in conductance. E. Voltage clamp recordings in RFP-positive hippocampal neurons revealed that GLP-1 either elicits an inward current (left panel) as in PVN and BNST, or an outward current. F. Mean data from current clamp recordings (left panel) demonstrating that GLP-1 causes a 10–15 mV depolarisation in most neurons tested, and a hyperpolarisation in individual hippocampal cells. Mean data from voltage clamp recordings in the right panel demonstrate the inward current of 10–20 pA amplitude elicited by 100 nM GLP-1 underlying the depolarisation in current clamp. n-numbers are given below the bars. **: p < 0.01.
© Copyright Policy - CC BY
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4588458&req=5

fig7: GLP-1 elicits electrical responses in GLP-1R-Cre positive cells in various brain regions. A, B. Voltage-clamp recording from an RFP-positive PVN neuron. Bath application of 100 nM GLP-1 elicited a small inward current at a holding potential of −50 mV accompanied by an increase in whole-cell conductance. These effects reversed upon washout of GLP-1 from the bath solution. C, D. Similarly, in a BNST neuron bath application of 100 nM GLP-1 elicits an inward current accompanied by an increase in conductance. E. Voltage clamp recordings in RFP-positive hippocampal neurons revealed that GLP-1 either elicits an inward current (left panel) as in PVN and BNST, or an outward current. F. Mean data from current clamp recordings (left panel) demonstrating that GLP-1 causes a 10–15 mV depolarisation in most neurons tested, and a hyperpolarisation in individual hippocampal cells. Mean data from voltage clamp recordings in the right panel demonstrate the inward current of 10–20 pA amplitude elicited by 100 nM GLP-1 underlying the depolarisation in current clamp. n-numbers are given below the bars. **: p < 0.01.
Mentions: Seven red-fluorescent PVN neurons were recorded in voltage clamp at a holding potential of −50 mV, and five cells were recorded in current-clamp mode. The mean resting potential in current-clamp mode was −57 ± 2 mV (n = 5). Bath-application of 100 nM GLP-1 caused a depolarisation by 14 ± 1 mV in all five cells that reversed upon washout of the drug. In voltage clamp GLP-1 application induced a reversible inward current of 16 ± 3 pA (n = 7) that was accompanied by a decrease in membrane resistance from 1.1 ± 0.2 GΩ to 0.8 ± 0.1 GΩ (Figure 7A–C, F).

Bottom Line: Large numbers of eYFP or tdRFP immunoreactive cells were found in the circumventricular organs, amygdala, hypothalamic nuclei and the ventrolateral medulla.However, tdRFP positive neurons were also found in areas without preproglucagon-neuronal projections like hippocampus and cortex.GLP-1R expression was confirmed in whole-cell recordings from BNST, hippocampus and PVN, where 100 nM GLP-1 elicited a reversible inward current or depolarisation.

View Article: PubMed Central - PubMed

Affiliation: Centre for Cardiovascular and Metabolic Neuroscience, Department of Neuroscience, Physiology & Pharmacology, University College London, London, WC1E 6BT, UK.

ABSTRACT

Objective: Although Glucagon-like peptide 1 is a key regulator of energy metabolism and food intake, the precise location of GLP-1 receptors and the physiological relevance of certain populations is debatable. This study investigated the novel GLP-1R-Cre mouse as a functional tool to address this question.

Methods: Mice expressing Cre-recombinase under the Glp1r promoter were crossed with either a ROSA26 eYFP or tdRFP reporter strain to identify GLP-1R expressing cells. Patch-clamp recordings were performed on tdRFP-positive neurons in acute coronal brain slices from adult mice and selective targeting of GLP-1R cells in vivo was achieved using viral gene delivery.

Results: Large numbers of eYFP or tdRFP immunoreactive cells were found in the circumventricular organs, amygdala, hypothalamic nuclei and the ventrolateral medulla. Smaller numbers were observed in the nucleus of the solitary tract and the thalamic paraventricular nucleus. However, tdRFP positive neurons were also found in areas without preproglucagon-neuronal projections like hippocampus and cortex. GLP-1R cells were not immunoreactive for GFAP or parvalbumin although some were catecholaminergic. GLP-1R expression was confirmed in whole-cell recordings from BNST, hippocampus and PVN, where 100 nM GLP-1 elicited a reversible inward current or depolarisation. Additionally, a unilateral stereotaxic injection of a cre-dependent AAV into the PVN demonstrated that tdRFP-positive cells express cre-recombinase facilitating virally-mediated eYFP expression.

Conclusions: This study is a comprehensive description and phenotypic analysis of GLP-1R expression in the mouse CNS. We demonstrate the power of combining the GLP-1R-CRE mouse with a virus to generate a selective molecular handle enabling future in vivo investigation as to their physiological importance.

No MeSH data available.


Related in: MedlinePlus