# Essential Roles of Heparan Sulfate Endosulfatase Sulf1 in Reward and Aversion Learning Through Distinct Dopamine D1 and D2 Receptor Pathways in Male Mice

## TL;DR

This study shows that Sulf1, an enzyme in the brain, is important for learning about rewards and avoiding unpleasant experiences through different dopamine pathways in mice.

## Contribution

The study reveals distinct roles of Sulf1 in dopamine D1 and D2 receptor pathways for reward and aversion learning in adult mice.

## Key findings

- Sulf1 knockout mice showed impaired reward learning in the cocaine-induced CPP test and aversion learning in the IA test.
- Conditional Sulf1 disruption in D1-expressing cells impaired reward learning, while disruption in D2-expressing cells impaired aversion learning.
- Sulf1 is essential for proper function of NAc circuitry in reward and aversion behaviors.

## Abstract

Sulf1 and Sulf2 are extracellular sulfatases that remove 6‐O‐sulfate from heparan sulfate and thereby regulate cell signaling. Previous studies have revealed that Sulf1/Sulf2 double knockout (KO) mice had defects in differentiation and axon guidance during development, but their functional roles in the adult brain remain largely unknown. We recently found that Sulf1 mRNA is highly expressed in the nucleus accumbens (NAc) shell and that Sulf1 expression is detected in both types of medium spiny neurons expressing dopamine D1 or D2 receptors. Moreover, we found that Sulf1 KO led to changes in membrane excitability and excitatory synaptic transmission in medium spiny neurons of the NAc in adult mice. These findings suggest possible roles of Sulf1 in the functions of NAc circuitry. To address this question, we performed behavioral tests using Sulf1 KO mice. We found that constitutive Sulf1 KO mice showed impairment in both the cocaine‐induced conditioned place preference (CPP) test and inhibitory avoidance (IA) test. Next, to examine which cell types the Sulf1 gene is required for, we generated Sulf1 floxed mice by means of CRISPR‐Cas9‐mediated genome editing and mated them with mice expressing Cre recombinase under a promoter for either the dopamine D1 or D2 receptor‐encoding genes. Sulf1 conditional knockout (cKO) in cells expressing dopamine D1 receptors led to impairment only in the CPP test, whereas Sulf1 cKO in D2 receptor‐expressing cells resulted in impairment only in the IA test. These results demonstrate that Sulf1 is required for both reward and aversion learning, and that the D1‐ and D2‐pathways distinctly regulate these functions. The present study suggests that Sulf1 is essential for neuronal functions and behavioral control in the adult brain.

In the nucleus accumbens (NAc), a heparan sulfate endosulfatase Sulf1 is expressed in both dopamine D1 receptor (Drd1) expressing and Drd2‐expressing medium spiny neurons (D1‐MSNs and D2‐MSNs, respectively). Sulf1 knockout (KO) mice showed impairment in both the cocaine‐induced conditioned place preference (CPP) test and inhibitory avoidance (IA) test. Conditional Sulf1 disruption in Drd1‐ and Drd2‐expressing cells led to impairment in the CPP test and IA test, respectively. These results demonstrated that Sulf1 is required for both reward and aversion learning, and that the D1‐ and D2‐pathways distinctly regulate these functions.