G Protein-coupled receptors (GPCRs) are transmembrane proteins that represent a highly important class of drug targets, being the primary target of about 35% of all marketed drugs. A prime family of GPCRs is the family of histamine receptors consisting of four distinct members named histamine H1 – H4 receptor (H1R-H4R). For further study of the role of localized activation of histamine receptors in pathophysiology, the development of photoswitchable ligands may be of aid. Photoswitchable ligands are ligands that combine pharmacophore features of known histamine receptor ligands with a photoswitchable moiety. A number of photoswitchable moieties have been discovered over the course of history. However, we focused on the azobenzene moiety throughout this thesis due to its robustness and structural simplicity. Moreover, when illuminated the azobenzene moiety isomerizes from trans to cis, considerably altering the shape and polarity. One may imagine, that these difference in shape, end-to-end distance and polarity, the trans isomer and cis isomer can have different pharmacological outcomes. These different pharmacological outcomes can present itself in differences in binding affinity, efficacy and potency for the histamine receptor of interest between the two isomers. In Chapter 2 the photopharmacology concept was applied to the H3R where key compounds VUF14862 and VUF14738 were identified, showing respectively a 11.2-fold decrease or 13.5-fold increase in affinity upon illumination. In two-electrode voltage clamp (TEVC) experiments using Xenopus laevis oocytes expressing H3R and G protein-coupled inward-rectifying potassium channels (GIRK) it was shown that VUF14862 and VUF14738 were able to reversibly modulate their affinity for H3R upon illumination on a second-timescale. In Chapter 3, VUF15000 was identified as a key compound based on its high absolute potency as well as its full agonism in [35S]-GTPS assays for the H3R. When VUF15000 was applied in TEVC experiments it was able to show reversible activation of the H3R on a second-timescale. In Chapter 4, photoswitchable antagonists for the H4R were developed. Photoswitchable antagonist VUF16822 was identified as a key compound, showing >186-fold increase in H4R affinity upon illumination. The scaffold shows an appreciable red-shift of its isomerization wavelength, allowing for illumination with wavelengths in the visible range. In Chapter 5, VUF16129 was identified as an efficient H4R agonist showing a 15-fold difference in potency between the trans and PSS cis samples whilst acting as a nearly full agonist in both samples. The compound was further evaluated in a mouse itch model which showed that the trans isomer elicits an H4R-mediated itch response. Gratifyingly, when 365 nm light is applied to the mouse the H4R-mediated itch is strongly decreased, which is reversible using 440 nm light. In Chapter 6, three strategies were used to develop photoswitchable H1R antagonists which identified VUF16828 as a lead compound. The compound shows a nanomolar affinity for H1R, but only a 5-fold increase in affinity upon illumination. All in all, throughout this thesis we present 6 key compounds which serve as novel tool compounds for investigating pharmacology of histamine receptors through specific activation with spatiotemporal precision. As presented in various chapters, these photoswitches allow for precise control over pathophysiological processes like itch and pain perception. Ultimately, the photoswitchable ligands developed have illuminated new research avenues in the field of histamine (photo)pharmacology.