Dopaminergic inhibition of gonadotropin-releasing hormone neurons in the cichlid fish Astatotilapia burtoni

Abstract

Dopamine regulates reproduction in part by modulating neuronal activity within the hypothalamic-pituitary-gonadal (HPG) axis. Previous studies suggested numerous mechanisms by which dopamine exerts inhibitory control over the HPG axis, ultimately changing the levels of sex steroids that regulate reproductive behaviors. However, it is not known whether these mechanisms are conserved across vertebrate species. In particular, it is unknown whether mechanisms underlying dopaminergic control of reproduction are shared between mammals and teleost fish. In mammals, dopamine directly inhibits gonadotropin-releasing hormone (GnRH1) hypothalamic neurons, the gatekeepers for activation of the HPG axis. Here, we demonstrate, for the first time in teleost fish, dopaminergic control of GnRH1 neurons via direct dopamine type-2-like receptor (D2R)-mediated inhibition within the hypothalamus. These results suggest that direct dopaminergic control of GnRH1 neurons via interactions in the hypothalamus is not exclusive to tetrapod reproductive control, but is likely conserved across vertebrate species.

Keywords: Dopamine; GnRH; HPG axis; Hypothalamus; Reproduction.

Fig. 1.

Presence of dopaminergic processes in the preoptic area (POA). Upper row: maximum projection confocal image of eGFP-labeled gonadotropin-releasing hormone (GnRH1) neurons (green) surrounded by anti-tyrosine hydroxylase (TH)-immunostained processes and cell bodies (magenta). Staining replicated across three fish. Dorsal is up. Scale bar, 100 µm. Lower row: high magnification maximum projection confocal image showing close apposition of eGFP-expressing GnRH1 neurons and a TH-immunostained neuron (arrowhead). Scale bar, 10 µm.

Fig. 2.

Dopamine and dopamine type-2-like receptor (D2R) agonists hyperpolarize GnRH1 neurons in the POA. (A) Example intracellular recording from a tonically firing GnRH1 neuron; bath application of dopamine (1 µmol l⁻¹; gray bar) hyperpolarizes the resting membrane potential (V_m) below threshold. (B) Bath application of 1, 10 or 100 µmol l⁻¹ dopamine hyperpolarizes GnRH1 neurons (N=6 neurons, P=0.03, two-tailed Wilcoxon t-test). Neurons were recorded from brain slices collected from five fish. Values are the mean steady-state voltage response for each neuron. The dashed line indicates no change after drug application (line of unity). The crosshair represents the median effect (center) for the population, with interquartile range (IQR). (C) Example intracellular recording from a GnRH1 neuron. Bath application of quinpirole (gray bar), a D2R-specific agonist, drives neuronal hyperpolarization. (D) Summary of the effect of quinpirole application on female and male GnRH1 neurons (N=16 neurons, P=0.0005, two-tailed Wilcoxon t-test). Each neuron was recorded from a unique brain slice, collected from 13 fish. Conventions as for B. (E) Bath application of SKF81297 (gray bar), a D1 receptor-specific agonist, does not elicit any change in resting membrane potential in an intracellular recording from a representative GnRH1 neuron. (F) Summary of the effect of SKF81297 application on female and male GnRH1 neurons (N=10 neurons, P=0.32, two-tailed Wilcoxon t-test). Each neuron was recorded from a unique brain slice, collected from eight fish. Conventions as for B.

Fig. 3.

Mechanisms underlying D2R-mediated hyperpolarization of GnRH1 neurons. (A) Representative intracellular recording from a GnRH1 neuron. Quinpirole (gray bar)-induced membrane hyperpolarization is not abolished by pre-treatment with the synaptic transmission blocker CdCl₂. (B) Across the population of recorded neurons, synaptic blockers (CdCl₂ and tetrodotoxin, TTX) did not affect membrane hyperpolarization elicited by quinpirole application (N=6 neurons, P=0.03, two-tailed Wilcoxon t-test). Neurons were recorded from brain slices collected from four fish. The cross-hair represents the median effect (center), with IQR. (C) Example plot showing membrane potential changes induced by current injection in a GnRH1 neuron before (left) and after (right) activation of D2Rs via quinpirole bath application. Traces represent sequential injection of current steps, from −100 to 0 pA. (D) Summary of the effect of quinpirole on the voltage–current (V–I) relationship in GnRH1 neurons. Symbols represent the mean current-evoked change in membrane potential. Lines represent linear regressions of the plotted population averages (N=14 neurons, P<0.001, ANCOVA). Each neuron was recorded from a unique brain slice, collected from 11 fish.