Journal: Acta Neuropathologica Communications

Article Title: Cellular and subcellular localization of Rab10 and phospho-T73 Rab10 in the mouse and human brain

doi: 10.1186/s40478-023-01704-9

Figure Lengend Snippet: Rab10 and pRab10 immunofluorescence and confocal microscope images of C57BL/6J WT mouse brain striatum. a Rab10 (green) expression in DARPP32 (red) positive SPNs indicated by arrows, and parvalbumin (blue) positive interneurons indicated by arrowheads. Scale bar = 50 µm. b pRab10 (green) expression in DARPP32 (red) positive SPNs indicated by arrows and parvalbumin (blue) positive interneurons indicated by arrowheads. Scale bar = 50 µm. c A representative image shows Rab10 (green) expression in ChAT (red) positive interneurons in merged and higher magnification images indicated by arrows. Scale bar = 50, 10 µm. d A representative image shows pRab10 (green) expression in ChAT (red) positive interneurons in merged and higher magnification images indicated by arrows. Scale bar = 50, 10 µm. (n = 3 biologically independent mouse brain samples)

Article Snippet: Rab10 (4262S, Cell Signaling Technology) for immunoblot, Rab10 (ab237703, Abcam) for immunofluorescence, pRab10 for immunoblot (ab230261, Abcam), pRab10 for immunofluorescence (ab241060, Abcam), Hsc70 (ab19136, Abcam), α-Tubulin (SC23948, Santa Cruz Biotechnology), parvalbumin, (NBP2-50036, NovusBio), NeuN (MAB377, Millipore), SATB2 (ab51502 Abcam), calretinin (MAB1568, Millipore), DARPP32 (MAB4230, R&D Systems), ChAT (NBP2-46620, NovusBio), DAT loop (6-8D6 Santa Cruz Biotechnology), tyrosine hydroxylase (TH) (ab76442, Abcam), CD68 (NBP2-33337SS, NovusBio), GFAP (AB5541, Millipore), Olig2 (MABN50 Millipore), KDEL receptor (sc-58774, Santa Cruz Biotechnology), TGN46 (MA3-063, ThermoFisher), LAMP1 (1D4B, DHSB), EEA1 (NBP2-36568, NovusBio), α-Synuclein (610786, BD Transduction Lab), Synuclein (ab51252, Abcam), VAMP2 (104 211, Synaptic Systems), Homer1 (160 006 Synaptic Systems), Rab8a (ab188574).

Techniques: Immunofluorescence, Microscopy, Expressing

Journal: Cancers

Article Title: The Prostate Cancer Therapy Enzalutamide Compared with Abiraterone Acetate/Prednisone Impacts Motivation for Exploration, Spatial Learning and Alters Dopaminergic Transmission in Aged Castrated Mice

doi: 10.3390/cancers13143518

Figure Lengend Snippet: ENZ modulates midbrain dopaminergic release to striatum. Note. ( A ) Schematic representation of SNpC dopaminergic projections to striatum and consequent striatal neuronal activation. Immunofluorescence of TH + -DA striatal projections of ENZ-, AAP- and vehicle-treated mice (seen on magnification of squared areas). Scale bar: 100 μm. On the right, Whisker boxes represent TH + fibers intensity and the number of P-DARPP-32 + cells in the striatum of ENZ- and AAP-treated mice compared with respective vehicles. Statistical quantifications were performed using Mann–Whitney test. Data are represented as box and whiskers of mean ± SEM ( n = 4 mice), * p < 0.05. Scale bar: 100 μm. ( B ) Representative images of Dopamine cAMP-Regulated Neuronal Phosphoprotein P-DARPP-32 + —(green), DA receptor 1 + —(DR1, cyan), TH + —(red) positive fibers and DAPI (blue) immunoreactivities in striatum of ENZ-, AAP- and Vehicle-treated mice. Scale bar: 25 μm and 100 μm. AAP, abiraterone acetate-prednisone; DA, dopamine; DAPI, 4′,6-diamidino-2-phenylindol; ENZ, enzalutamide; P-DARPP-32, Phosphorylated form of Dopamine cAMP-Regulated Neuronal Phosphoprotein; DR1, DA receptor 1; SNpc, substantia nigra pars compacta; TH, Tyrosine hydroxylase; vHp, ventral hippocampus.

Article Snippet: The following primary antibodies and the corresponding dilutions were used as followed: anti-GFAP (Goat, Abcam, Cambridge, UK, AB53554,1:1000) or anti-GFAP (Rabbit, Dako, Agilent, Santa Clara, CA, USA, Z0334), anti-AR (Rabbit, Abcam AB74272, 1:300), anti-BDNF (Goat, St John’s Lab, London, UK, STJ73299, 1:1000), anti-TH (Goat, Abcam AB101853) or anti-TH (chicken, Abcam AB76442,1:500), anti-Phospho-DARPP-32 (P-DARPP-32, Rabbit, Bioss, Boston, MA, USA, bs-3118R, 1:200), anti-BrdU (Sheep, Abcam AB1893, 1:200), anti-NeuN (Rabbit, Abcam AB104225) or anti-NeuN (Chicken, MyBiosource MBS837654, 1:1000), anti-c-fos (Rabbit, Abcam AB190289), anti-DCX (Guinea pig, Millipore, Burlington, MA, USA), anti-D1R (Rabbit, Abcam, AB20066, 1:200) or anti-CYP17A1 (Rabbit, LifeSpan Biosciences Inc.,Seattle, WA, USA, LS-C352100, 1:100).

Techniques: Activation Assay, Immunofluorescence, Whisker Assay, MANN-WHITNEY

Journal: Cancers

Article Title: The Prostate Cancer Therapy Enzalutamide Compared with Abiraterone Acetate/Prednisone Impacts Motivation for Exploration, Spatial Learning and Alters Dopaminergic Transmission in Aged Castrated Mice

doi: 10.3390/cancers13143518

Figure Lengend Snippet: ENZ modulates midbrain dopaminergic release to ventral hippocampus. Note. ( A ) Schematic of dopaminergic projections to ventral hippocampus (vHp) from VTA and consequent neuronal activation. Immunofluorescence of TH + -DA projections likely arising from VTA and projecting to vHp of ENZ-, AAP- and vehicle-treated mice (seen on magnification of squared areas). Scale bar: 100 μm. On the right, Whisker boxes represent the intensity of TH + fibers and number of P-DARPP-32 + cells in the vHp (right panel) of ENZ- and AAP-treated mice compared with respective vehicles. Statistical quantifications were performed using Mann–Whitney test. Data are represented as box and whiskers of mean ±SEM ( n = 4 mice), * p < 0.05. Scale bar: 100 μm. ( B ) Representative images of P-DARPP-32 + —(green), DR1 + —(cyan), TH + —(red) positive fibers and DAPI (blue) immunoreactivities in vHp of ENZ-, AAP- and Vehicle-treated mice. Scale bar: 25 μm and 100 μm. ( C ) Representative example of the immediate early gene (c-fos, green) and NeuN (red) immunoreactivities in the DG, CA3 and CA1 regions of vHp of ENZ-, AAP- and vehicle-treated mice. The boxed areas show DAPI (blue) immunoreactivity. On the left, whisker boxes represent the number of c-fos + cells in DG, CA3 and CA1 hippocampal areas of ENZ- and AAP-treated mice compared with respective vehicles. Statistical quantifications were performed using Mann–Whitney test. Data are represented as box and whiskers and mean ± SEM ( n = 4 mice), * p < 0.05, ** p ≤ 0.01. Scale bar: 100 μm. AAP, abiraterone acetate-prednisone; CA1, cornus ammonis 1; CA3, cornus ammonis 3; c-fos, cellular oncogene c-fos; DA, dopamine; DAPI, 4′,6-diamidino-2-phenylindol; DG, dentate gyrus; ENZ, enzalutamide; NeuN, Neuronal Nuclei Antigen; P-DARPP-32, Phosphorylated form of Dopamine cAMP-Regulated Neuronal Phosphoprotein; DR1, DA receptor 1; TH, Tyrosine hydroxylase; vHp, ventral hippocampus; VTA, ventral tegmental area.

Article Snippet: The following primary antibodies and the corresponding dilutions were used as followed: anti-GFAP (Goat, Abcam, Cambridge, UK, AB53554,1:1000) or anti-GFAP (Rabbit, Dako, Agilent, Santa Clara, CA, USA, Z0334), anti-AR (Rabbit, Abcam AB74272, 1:300), anti-BDNF (Goat, St John’s Lab, London, UK, STJ73299, 1:1000), anti-TH (Goat, Abcam AB101853) or anti-TH (chicken, Abcam AB76442,1:500), anti-Phospho-DARPP-32 (P-DARPP-32, Rabbit, Bioss, Boston, MA, USA, bs-3118R, 1:200), anti-BrdU (Sheep, Abcam AB1893, 1:200), anti-NeuN (Rabbit, Abcam AB104225) or anti-NeuN (Chicken, MyBiosource MBS837654, 1:1000), anti-c-fos (Rabbit, Abcam AB190289), anti-DCX (Guinea pig, Millipore, Burlington, MA, USA), anti-D1R (Rabbit, Abcam, AB20066, 1:200) or anti-CYP17A1 (Rabbit, LifeSpan Biosciences Inc.,Seattle, WA, USA, LS-C352100, 1:100).

Techniques: Activation Assay, Immunofluorescence, Whisker Assay, MANN-WHITNEY

Journal: Cancers

Article Title: The Prostate Cancer Therapy Enzalutamide Compared with Abiraterone Acetate/Prednisone Impacts Motivation for Exploration, Spatial Learning and Alters Dopaminergic Transmission in Aged Castrated Mice

doi: 10.3390/cancers13143518

Figure Lengend Snippet: Schematic illustration of the central action of novel generation hormone therapy targeting the androgen signaling axis, abiraterone acetate+prednisone (AAP) and enzalutamide (ENZ) in aged castrated mice. Note. ( A ) ENZ which is able to cross the blood–brain barrier (BBB) can bind and inhibit ARs expressed in peripheral and brain organs. More specifically, CYP17A1 and AR are localized in both SNpc and VTA neurons in aged castrated mouse brains. ENZ leads to reduced P-DARPP-32 activated signaling in mature neurons expressing DR1 in the striatum through the nigrostriatal pathway and in the vHP via part of the mesolimbocortical pathway. This inhibition of DA-expressing afferents likely accounts for diminished locomotor activity and exploration behavior. ENZ also depressed mature neuron activity evidenced by c-fos expression in dHP potentially responsible for an altered spatial cognitive performance. ( B ) AAP is a prodrug, generating abiraterone selective for CYP17A1 enzyme activity, thus inhibiting androgen biosynthesis in the testis and the adrenal gland, but potentially in the VTA and SNpc. AA may have the ability to cross the BBB. In order to avoid cortisol deficiency, the treatment is combined with chronic use of prednisone (P) sufficient to activate the HPA-related feedback loop and to reduce the plasma ACTH level allowing physiological cortisol production in patients. CYP17A1 and AR are expressed in the dHP, and here the AAP effect on proliferation of precursor cells and inhibition of BDNF production without modification of mature neurons would hypothetically involve glucocorticoids receptors. In the vHP, the AAP-evoked reduction of c-Fos in mature neurons can sustain the delay in efficiency in spatial learning acquisition. AA, abiraterone acetate; AAP, abiraterone acetate-prednisone; ACTH, adrenocorticotropic hormone; AR, androgen receptor; BBB, blood–brain barrier; BDNF, Brain-derived neurotrophic factor; c-fos, cellular oncogene c-fos; CYP17A1, Cytochrome P450 Family 17 Subfamily A Member 1; DA, dopamine; dHP, dorsal hippocampus; ENZ, enzalutamide; HPA, hypothalamic–pituitary–adrenal; P, prednisone; P-DARPP-32, Phosphorylated form of Dopamine cAMP-Regulated Neuronal Phosphoprotein; DR1, DA receptor 1; SNpc, substantia nigra pars compacta; vHP, ventral hippocampus; VTA, ventral tegmental area.

Article Snippet: The following primary antibodies and the corresponding dilutions were used as followed: anti-GFAP (Goat, Abcam, Cambridge, UK, AB53554,1:1000) or anti-GFAP (Rabbit, Dako, Agilent, Santa Clara, CA, USA, Z0334), anti-AR (Rabbit, Abcam AB74272, 1:300), anti-BDNF (Goat, St John’s Lab, London, UK, STJ73299, 1:1000), anti-TH (Goat, Abcam AB101853) or anti-TH (chicken, Abcam AB76442,1:500), anti-Phospho-DARPP-32 (P-DARPP-32, Rabbit, Bioss, Boston, MA, USA, bs-3118R, 1:200), anti-BrdU (Sheep, Abcam AB1893, 1:200), anti-NeuN (Rabbit, Abcam AB104225) or anti-NeuN (Chicken, MyBiosource MBS837654, 1:1000), anti-c-fos (Rabbit, Abcam AB190289), anti-DCX (Guinea pig, Millipore, Burlington, MA, USA), anti-D1R (Rabbit, Abcam, AB20066, 1:200) or anti-CYP17A1 (Rabbit, LifeSpan Biosciences Inc.,Seattle, WA, USA, LS-C352100, 1:100).

Techniques: Expressing, Inhibition, Activity Assay, Modification, Derivative Assay