Background: Botulinum neurotoxins (BoNTs) are used to treat spastic disorders. Depending on muscle size, one or multiple injections are recommended according to labels to target neuromuscular junctions (NMJ). However, information about NMJ distribution and number in muscles, as well as expression of receptors and molecular targets of toxins is scarce in human and animal models. Methods: Seven muscles from adult rats were used to identify expression of BoNT receptors and SNAREs using immunohistochemistry (IHC), and fluorescent alpha-Bungarotoxin combined to light-sheet microscopy used to determine their distribution. Results: The location, number, and density of NMJ were muscle specific and mostly dependent on the type of pennation (myofiber orientation). In the Flexor Digitorum Brevis (a very small muscle) NMJ were as numerous as in the Gastrocnemius lateralis. A strong expression of SV2C, Synaptotagmin 2, SNAP25 and VAMP1 were observed in all muscles, and SV2A, Synaptotagmin 1 and VAMP2 were never detected. Conclusion: This work highlights the specific distribution of NMJ in muscles which seems to depend on the type of pennation. Detailed observation of myofibers organization might help clinicians to better evaluate the location of NMJ in humans; the molecular phenotyping of NMJ will contribute to better integrate the rat model into research of BoNT therapeutics.

Limited experiments have compared the treatment effects of repetitive cycles of radiolabelled somatostatin (SST) analogues. In vitro and in vivo experiments were conducted in an AR42J cancer cell model, comparing the antagonist [¹⁷⁷Lu]Lu-satoreotide tetraxetan with the agonist [¹⁷⁷Lu]Lu-DOTA-TATE in terms of their binding properties, biodistribution, anti-tumour activity and toxicity. Histopathological and immunohistochemical examinations were performed at different timepoints. In the in vitro assays, [¹⁷⁷Lu]Lu-satoreotide tetraxetan recognised twice as many SST₂ binding sites as [¹⁷⁷Lu]Lu-DOTA-TATE. In mice treated once a week for four consecutive weeks, [¹⁷⁷Lu]Lu-satoreotide tetraxetan (15 MBq) revealed a significantly greater median time taken to reach a tumour volume of 850 mm³ (68 days) compared to [¹⁷⁷Lu]Lu-DOTA-TATE at 15 MBq (43 days) or 30 MBq (48 days). This was associated with a higher tumour uptake, enhanced DNA damage and no or mild effects on body weight, haematological toxicity, or renal toxicity with [¹⁷⁷Lu]Lu-satoreotide tetraxetan (15 MBq). At the end of the study, complete tumour senescence was noted in 20% of animals treated with [¹⁷⁷Lu]Lu-satoreotide tetraxetan, in 13% of those treated with [¹⁷⁷Lu]Lu-DOTA-TATE at 30 MBq, and in none of those treated with [¹⁷⁷Lu]Lu-DOTA-TATE at 15 MBq. In conclusion, repeated administrations of [¹⁷⁷Lu]Lu-satoreotide tetraxetan were able to potentiate peptide receptor radionuclide therapy with a higher tumour uptake, longer median survival, and enhanced DNA damage, with a favourable efficacy/safety profile compared to [¹⁷⁷Lu]Lu-DOTA-TATE.

Botulinum neurotoxins E (BoNT/E) and A (BoNT/A) act by cleaving Synaptosome-Associated Protein 25 (SNAP25) at two different C-terminal sites, but they display very distinct durations of action, BoNT/E being short acting and BoNT/A long acting. We investigated the duration of action, spread and neuronal transport of BoNT/E (6.5 ng/kg) and BoNT/A (125 pg/kg) after single intramuscular administrations of high equivalent efficacious doses, in rats, over a 30- or 75-day periods, respectively. To achieve this, we used (i) digit abduction score assay, (ii) immunohistochemistry for SNAP25 (N-ter part; SNAP25^N-ter and C-ter part; SNAP25^C-ter) and its cleavage sites (cleaved SNAP25; c-SNAP25^E and c-SNAP25^A) and (iii) muscular changes in histopathology evaluation. Combined in vivo observation and immunohistochemistry analysis revealed that, compared to BoNT/A, BoNT/E induces minimal muscular changes, possesses a lower duration of action, a reduced ability to spread and a decreased capacity to be transported to the lumbar spinal cord. Interestingly, SNAP25^C-ter completely disappeared for both toxins during the peak of efficacy, suggesting that the persistence of toxin effects is driven by the persistence of proteases in tissues. These data unveil some new molecular mechanisms of action of the short-acting BoNT/E and long-acting BoNT/A, and reinforce their overall safety profiles.