Dopamine Axon Targeting in the Nucleus Accumbens in Adolescence Requires Netrin-1

Abstract

The fine arrangement of neuronal connectivity during development involves the coordinated action of guidance cues and their receptors. In adolescence, the dopamine circuitry is still developing, with mesolimbic dopamine axons undergoing target-recognition events in the nucleus accumbens (NAcc), while mesocortical projections continue to grow toward the prefrontal cortex (PFC) until adulthood. This segregation of mesolimbic versus mesocortical dopamine pathways is mediated by the guidance cue receptor DCC, which signals dopamine axons intended to innervate the NAcc to recognize this region as their final target. Whether DCC-dependent mesolimbic dopamine axon targeting in adolescence requires the action of its ligand, Netrin-1, is unknown. Here we combined shRNA strategies, quantitative analysis of pre- and post-synaptic markers of neuronal connectivity, and pharmacological manipulations to address this question. Similar to DCC levels in the ventral tegmental area, Netrin-1 expression in the NAcc is dynamic across postnatal life, transitioning from high to low expression across adolescence. Silencing Netrin-1 in the NAcc in adolescence results in an increase in the expanse of the dopamine input to the PFC in adulthood, with a corresponding increase in the number of presynaptic dopamine sites. This manipulation also results in altered dendritic spine density and morphology of medium spiny neurons in the NAcc in adulthood and in reduced sensitivity to the behavioral activating effects of the stimulant drug of abuse, amphetamine. These cellular and behavioral effects mirror those induced by Dcc haploinsufficiency within dopamine neurons in adolescence. Dopamine targeting in adolescence requires the complementary interaction between DCC receptors in mesolimbic dopamine axons and Netrin-1 in the NAcc. Factors regulating either DCC or Netrin-1 in adolescence can disrupt mesocorticolimbic dopamine development, rendering vulnerability or protection to phenotypes associated with psychiatric disorders.

Unique Effects of Social Defeat Stress in Adolescent Male Mice on the Netrin-1/DCC Pathway, Prefrontal Cortex Dopamine and Cognition

“…2 B ). Since the complementary actions of DCC receptors on axons of VTA DA neurons and Netrin-1 in the NAcc dictate the segregation of mesolimbic and mesocortical projections ( Manitt et al, 2013 ; Reynolds et al, 2018 ; Cuesta et al, 2020a ), the downregulation of VTA Dcc mRNA in resilient mice suggests targeting errors of mesolimbic DA axons and concomitant changes in adult PFC DA connectivity. The upregulation of NAcc Netrin-1, in susceptible mice, suggests a compensatory effect.…”

“…As previously ( Torres-Berrío et al, 2017 ; Cuesta et al, 2020a ), protein samples (20 μg, n = 25 adolescents and n = 24 adults) were separated on a 10% SDS-PAGE and transferred to a PVDF membrane which was incubated overnight at 4°C with antibodies against Netrin-1 (Abcam, catalog #126729) and α-tubulin (Cell Signaling, catalog #2144S) for loading control. Four biological samples were included per experimental condition.…”

“…Mesocorticolimbic DA axons reach the nucleus accumbens (NAcc) by early adolescence, and while some remain there (mesolimbic system), others (mesocortical system) continue growing to the PFC until early adulthood ( Reynolds et al, 2018 ). The proper segregation of mesolimbic and mesocortical DA axons is regulated by the coordinated action of the guidance cue receptor DCC on DA axons and its ligand, Netrin-1, in the NAcc ( Hoops and Flores, 2017 ; Hoops et al, 2018 ; Reynolds et al, 2018 ; Cuesta et al, 2020a ).…”

DCC gene network in the prefrontal cortex is associated with total brain volume in childhood

“…[2][3][4][5][6][7][8][9][10][11] The gradual maturational trajectory of mesocor-tical dopamine connectivity -and of the PFC itself -is controlled by the guidance cue Netrin-1 and its receptor DCC, which determine the spatiotemporal targeting of growing axons. 10,12 Studies in rodents show that variations in Dcc gene expression lead to differences in cognitive flexibility, behavioural inhibition, vulnerability to stressinduced depression-like behaviours and altered sensitivity to drugs of abuse. 10,11,[13][14][15][16][17][18] In humans, an autosomal-dominant DCC mutation results in structural and functional alterations in the connectivity of mesocortical pathways, with an associated decrease in novelty-seeking behaviour and cigarette use.…”

“… 2 – 11 The gradual maturational trajectory of mesocortical dopamine connectivity — and of the PFC itself — is controlled by the guidance cue Netrin-1 and its receptor DCC, which determine the spatiotemporal targeting of growing axons. 10 , 12 Studies in rodents show that variations in Dcc gene expression lead to differences in cognitive flexibility, behavioural inhibition, vulnerability to stress-induced depression-like behaviours and altered sensitivity to drugs of abuse. 10 , 11 , 13 – 18 …”