Fig. 4: Downregulation of Arc mRNA levels in the LC, especially in TH+ but zif268- cells tracks the development of compulsive adjunctive polydipsic drinking behaviour.

The regulation of Arc mRNA levels within the LC by the expression of polydipsic drinking was not uniform across its different cell types and functional ensembles as shown by correlation matrices which track the relationship between the level of polydipsic drinking and (i) the total mRNA copies of zif268 and Arc in the LC and (ii) the representativity of various cell types characterised for their expression of the tyrosine hydroxylase, GFAP, zif268 and Arc, as well as any of their combinations. This dimensional analysis revealed that the negative relationship observed between the number of Arc mRNA copies in the LC and polydipsic drinking only emerged from session 8 onwards, exactly at the time when HD rats started to diverge from LD rats on their route towards the development of compulsive polydipsic drinking. This relationship was not observed for zif268, a distinction suggestive of a differential contribution of the two plasticity markers to the development of compulsive adjunctive behaviour and supported by a very similar pattern of correlation between polydipsic drinking and the percentage number of cells expressing Arc, but not zif268. Further analysis of these matrices revealed that the relationship between polydipsic drinking and Arc mRNA levels was only present in neurons (no correlation was found in GFAP+ cells), more precisely in TH+ neurons and only in those that do not belong to a zif268+ ensemble. Removing the water bottle at test, which prevented rats from expressing their adjunctive polydipsic behaviour, be it compulsive or not, disrupted the correlation with Arc mRNA levels, thereby revealing that the downregulation of Arc in TH+ zif268- cell ensembles tracks the expression of a compulsive adjunctive response.