Effects of the 5-HT1A receptor agonists buspirone and 8-OH-DPAT on pupil size in common marmosets.

As pupil size is affected by psychotropic drugs in all mammals, it has been used as a well-established clinical indicator for the preclinical and clinical development of novel drugs. It has been reported that activation of the serotonin (5-HT)1A receptor differently affects pupil response in rodents (mydriasis) and humans (miosis). Thus, it is important to establish a quantitative system for measuring pupil size using other species, such as nonhuman primates.

Ketamine-induced changes in connectivity of functional brain networks in awake female nonhuman primates: a translational functional imaging model.

Rationale: There is a significant interest in the NMDA-receptor antagonist ketamine due to its efficacy in treating depressive disorders and its induction of psychotic-like symptoms that make it a useful tool for modeling psychosis. Pharmacological MRI in awake nonhuman primates provides a highly translational model for studying the brain network dynamics involved in producing these drug effects.

Objective: The present study evaluated ketamine-induced changes in functional connectivity (FC) in awake rhesus monkeys.

Ketamine-induced brain activation in awake female nonhuman primates: a translational functional imaging model.

Rationale: There is significant interest in the NMDA receptor antagonist ketamine due to its efficacy in treating depressive disorders and its induction of psychotic-like symptoms that make it a useful tool for modeling psychosis.

Objective: The present study extends the successful development of an apparatus and methodology to conduct pharmacological MRI studies in awake rhesus monkeys in order to evaluate the CNS effects of ketamine.

Methods: Functional MRI scans were conducted in four awake adult female rhesus monkeys during sub-anesthetic intravenous (i.

Metabolism of 1alpha,25-dihydroxyvitamin D2 by human CYP24A1.

The metabolism of 1alpha,25-dihydroxyvitamin D2 (1alpha,25(OH)2D2) by human CYP24A1 was examined using the recombinant enzyme expressed in Escherichia coli cells. HPLC analysis revealed that human CYP24A1 produces at least 10 metabolites, while rat CYP24A1 produces only three metabolites, indicating a remarkable species-based difference in the CYP24A1-dependent metabolism of 1alpha,25(OH)2D2 between humans and rats. LC-MS analysis and periodate treatment of the metabolites strongly suggest that human CYP24A1 converts 1alpha,25(OH)2D2 to 1alpha,24,25,26(OH)4D2, 1alpha,24,25,28(OH)4D2, and 24-oxo-25,26,27-trinor-1alpha(OH)D2 via 1alpha,24,25(OH)3D2.

Kinetic studies of 25-hydroxy-19-nor-vitamin D3 and 1 alpha,25-dihydroxy-19-nor-vitamin D3 hydroxylation by CYP27B1 and CYP24A1.

Our previous study demonstrated that 25-hydroxy-19-nor-vitamin D(3) [25(OH)-19-nor-D(3)] inhibited the proliferation of immortalized noncancerous PZ-HPV-7 prostate cells similar to 1 alpha,25-dihydroxyvitamin D(3) [1 alpha,25(OH)(2)D(3)], suggesting that 25(OH)-19-nor-D(3) might be converted to 1 alpha,25-dihydroxy-19-nor-vitamin D(3) [1 alpha,25(OH)(2)-19-nor-D(3)] by CYP27B1 before exerting its antiproliferative activity. Using an in vitro cell-free model to study the kinetics of CYP27B1-dependent 1 alpha-hydroxylation of 25(OH)-19-nor-D(3) and 25-hydroxyvitamin D(3) [25(OH)D(3)] and CYP24A1-dependent hydroxylation of 1 alpha,25(OH)-19-nor-D(3) and 1 alpha,25(OH)(2)D(3), we found that k(cat)/K(m) for 1 alpha-hydroxylation of 25(OH)-19-nor-D(3) was less than 0.1% of that for 25(OH)D(3), and the k(cat)/K(m) value for 24-hydroxylation was not significantly different between 1 alpha,25(OH)(2)-19-nor-D(3) and 1 alpha,25(OH)(2)D(3).

Structure-function analysis of vitamin D 24-hydroxylase (CYP24A1) by site-directed mutagenesis: amino acid residues responsible for species-based difference of CYP24A1 between humans and rats.

Our previous studies revealed the species-based difference of CYP24A1-dependent vitamin D metabolism. Although human CYP24A1 catalyzes both C-23 and C-24 oxidation pathways, rat CYP24A1 shows almost no C-23 oxidation pathway. We tried to identify amino acid residues that cause the species-based difference by site-directed mutagenesis.

Interaction between mitochondrial CYP27B1 and adrenodoxin: role of arginine 458 of mouse CYP27B1.

A molecular modeling study of CYP27B1 suggests that Arg458 of mouse CYP27B1 is involved in interaction with adrenodoxin (ADX). Thus, we generated CYP27B1 mutants R458K and R458Q and revealed their enzymatic properties. Substrate-induced difference spectra and K(m) values for 1alpha-hydroxylation of 25(OH)D3 indicate that the replacement of Arg458 with Lys or Gln does not affect substrate binding.

Identification of the amino acid residue of CYP27B1 responsible for binding of 25-hydroxyvitamin D3 whose mutation causes vitamin D-dependent rickets type 1.

We previously reported the three-dimensional structure of human CYP27B1 (25-hydroxyvitamin D3 1alpha-hydroxylase) constructed by homology modeling. Using the three-dimensional model we studied the docking of the substrate, 25-hydroxyvitamin D3, into the substrate binding pocket of CYP27B1. In this study, we focused on the amino acid residues whose point mutations cause vitamin D-dependent rickets type 1, especially unconserved residues among mitochondrial CYPs such as Gln65 and Thr409.

Purification and characterization of mouse CYP27B1 overproduced by an Escherichia coli system coexpressing molecular chaperonins GroEL/ES.

The expression of mouse CYP27B1 in Escherichia coli has been dramatically enhanced by coexpression of GroEL/ES. To reveal the enzymatic properties of CYP27B1, we measured its hydroxylation activity toward vitamin D3 and 1alpha-hydroxyvitamin D3 (1alpha(OH)D3) in addition to the physiological substrate 25(OH)D3. Surprisingly, CYP27B1 converted vitamin D3 to 1alpha,25(OH)D3.