Lysosomes are key cellular catabolic centers that also perform fundamental metabolic, signaling and quality control functions. Lysosomes are not static and they respond dynamically to intra‐ and extracellular stimuli triggering changes in organelle numbers, size and position. Such physical changes have a strong impact on lysosomal activity ultimately influencing cellular homeostasis. In this review, we summarize the current knowledge on lysosomal size regulation, on its physiological role(s) and association to several disease conditions.

Therapeutic interest in augmentation of 5-hydroxytryptamine_2A (5-HT_2A) receptor signaling has been renewed by the effectiveness of psychedelic drugs in the treatment of various psychiatric conditions. In this study, we have further characterized the pharmacological properties of the recently developed 5-HT₂ receptor agonist N-2-hydroxybenzyl)-2,5-dimethoxy-4-cyanophenylethylamine (25CN-NBOH) and three structural analogs at recombinant 5-HT_2A, 5-HT_2B, and 5-HT_2C receptors and investigated the pharmacokinetic properties of the compound. 25CN-NBOH displayed robust 5-HT_2A selectivity in [³H]ketanserin/[³H]mesulergine, [³H]lysergic acid diethylamide and [³H]Cimbi-36 binding assays (K_i^2C/K_i^2A ratio range of 52–81; K_i^2B/K_i^2A ratio of 37). Moreover, in inositol phosphate and intracellular Ca²⁺ mobilization assays 25CN-NBOH exhibited 30- to 180-fold 5-HT_2A/5-HT_2C selectivities and 54-fold 5-HT_2A/5-HT_2B selectivity as measured by Δlog(R_max/EC₅₀) values. In an off-target screening 25CN-NBOH (10 μM) displayed either substantially weaker activity or inactivity at a plethora of other receptors, transporters, and kinases. In a toxicological screening, 25CN-NBOH (100 μM) displayed a benign acute cellular toxicological profile. 25CN-NBOH displayed high in vitro permeability (P_app = 29 × 10⁻⁶ cm/s) and low P-glycoprotein-mediated efflux in a conventional model of cellular transport barriers. In vivo, administration of 25CN-NBOH (3 mg/kg, s.c.) in C57BL/6 mice mice produced plasma and brain concentrations of the free (unbound) compound of ∼200 nM within 15 minutes, further supporting that 25CN-NBOH rapidly penetrates the blood-brain barrier and is not subjected to significant efflux. In conclusion, 25CN-NBOH appears to be a superior selective and brain-penetrant 5-HT_2A receptor agonist compared with (±)-2,5-dimethoxy-4-iodoamphetamine (DOI), and thus we propose that the compound could be a valuable tool for future investigations of physiologic functions mediated by this receptor.

Seizure threshold 2 (SZT2) is a component of the KICSTOR complex which, under catabolic conditions, functions as a negative regulator in the amino acid-sensing branch of mTORC1. Mutations in this gene cause a severe neurodevelopmental and epileptic encephalopathy whose main symptoms include epilepsy, intellectual disability, and macrocephaly. As SZT2 remains one of the least characterized regulators of mTORC1, in this work we performed a systematic interactome analysis under catabolic and anabolic conditions. Besides numerous mTORC1 and AMPK signaling components, we identified clusters of proteins related to autophagy, ciliogenesis regulation, neurogenesis, and neurodegenerative processes. Moreover, analysis of SZT2 ablated cells revealed increased mTORC1 signaling activation that could be reversed by Rapamycin or Torin treatments. Strikingly, SZT2 KO cells also exhibited higher levels of autophagic components, independent of the physiological conditions tested. These results are consistent with our interactome data, in which we detected an enriched pool of selective autophagy receptors/regulators. Moreover, preliminary analyses indicated that SZT2 alters ciliogenesis. Overall, the data presented form the basis to comprehensively investigate the physiological functions of SZT2 that could explain major molecular events in the pathophysiology of developmental and epileptic encephalopathy in patients with SZT2 mutations.