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The interest in the photochromism and functional applications of donor-acceptor Stenhouse adducts (DASAs) soared in recent years owing to their outstanding advantages and flexible design. However, their low solubility and irreversible conversion in aqueous solutions hampered exploring DASAs for biology and medicine. It is notably unknown whether the barbiturate electron acceptor group retains the pharmacological activity of drugs such as phenobarbital, which targets γ-aminobutyric acid (GABA)-type A receptors (GABAARs) in the brain. Here, we have developed the model compound DASA-barbital based on a scaffold of red-switching second-generation DASAs, and we demonstrate that it is active in GABAARs and alters the neuronal firing rate in a physiological medium at neutral pH. DASA-barbital can also be reversibly photoswitched in acidic aqueous solutions using cyclodextrin, an approved ingredient of drug formulations. These findings clarify the path toward the biological applications of DASAs and to exploit the versatility displayed in polymers and materials science.

Over the last decades, photopharmacology has gone far beyond its proof-of-concept stage to become a bona fide approach to study neural systems in vivo. Indeed, photopharmacological control has expanded over a wide range of endogenous targets, such as receptors, ion channels, transporters, kinases, lipids, and DNA transcription processes. In this review, we provide an overview of the recent progresses in the in vivo photopharmacological control of neuronal circuits and behavior. In particular, the use of small aquatic animals for the in vivo screening of photopharmacological compounds, the recent advances in optical modulation of complex behaviors in mice, and the development of adjacent techniques for light and drug delivery in vivo are described.

The interest in the photochromism and functional applications of donor-acceptor Stenhouse adducts (DASAs) soared in recent years, owing to their outstanding advantages and flexible design. However, their low solubility and irreversible conversion in aqueous solutions hampered exploring DASAs for biology and medicine. It is notably unknown whether the barbiturate electron acceptor group retains the pharmacological activity of drugs like phenobarbital, which targets γ-aminobutyric acid (GABA) type A receptors (GABAARs) in the brain. Here, we have developed the model compound DASA-barbital based on a scaffold of red-switching second-generation DASAs and we demonstrate that it is active in GABAARs and alters the neuronal firing rate in a physiological medium at neutral pH. DASA-barbital can also be reversibly photoswitched in acidic aqueous solutions using cyclodextrin, an approved ingredient of drug formulations. These findings clear the path towards the biological applications of DASAs and to exploit the versatility displayed in polymers and materials science.

Visual Abstract, Photopharmacology is a unique approach that through a combination of photochemistry methods and advanced life science techniques allows the study and control of specific biological processes, ranging from intracellular pathways to brain circuits. Recently, a first photochromic channel blocker of anion-selective GABAA receptors, the azobenzene-nitrazepam-based photochromic compound (Azo-NZ1), has been described. In the present study, using patch-clamp technique in heterologous system and in mice brain slices, site-directed mutagenesis and molecular modeling we provide evidence of the interaction of Azo-NZ1 with glycine receptors (GlyRs) and determine the molecular basis of this interaction. Glycinergic synaptic neurotransmission determines an important inhibitory drive in the vertebrate nervous system and plays a crucial role in the control of neuronal circuits in the spinal cord and brain stem. GlyRs are involved in locomotion, pain sensation, breathing, and auditory function, as well as in the development of such disorders as hyperekplexia, epilepsy, and autism. Here, we demonstrate that Azo-NZ1 blocks in a UV-dependent manner the activity of α2 GlyRs (GlyR2), while being barely active on α1 GlyRs (GlyR1). The site of Azo-NZ1 action is in the chloride-selective pore of GlyR at the 2’ position of transmembrane helix 2 and amino acids forming this site determine the difference in Azo-NZ1 blocking activity between GlyR2 and GlyR1. This subunit-specific modulation is also shown on motoneurons of brainstem slices from neonatal mice that switch during development from expressing “fetal” GlyR2 to “adult” GlyR1 receptors.

International audience; Glycine receptors (GlyRs) are indispensable for maintaining excitatory/inhibitory balance in neuronal circuits that control reflexes and rhythmic motor behaviors. Here we have developed Glyght, a GlyR ligand controlled with light. It is selective over other Cys-loop receptors, is active in vivo, and displays an allosteric mechanism of action. The photomanipulation of glycinergic neurotransmission opens new avenues to understanding inhibitory circuits in intact animals and to developing drug-based phototherapies.

Optogenetic and photopharmacological tools to manipulate neuronal inhibition have limited efficacy and reversibility. We report the design, synthesis, and biological evaluation of Fulgazepam, a fulgimide derivative of benzodiazepine that behaves as a pure potentiator of ionotropic γ‐aminobutyric acid receptors (GABAARs) and displays full and reversible photoswitching in vitro and in vivo. The compound enables high‐resolution studies of GABAergic neurotransmission, and phototherapies based on localized, acute, and reversible neuroinhibition., Fulgazepam: Herein, the design, synthesis, and biological evaluation of Fulgazepam, a fulgimide derivative of benzodiazepine that behaves as a pure potentiator of ionotropic γ‐aminobutyric acid receptors (GABAARs) and displays full and reversible photoswitching in vitro and in vivo, is reported.

Glycine receptors (GlyRs) belong to the family of ligand-gated cys-loop receptors and effectuate fast inhibitory neurotransmission in central nervous system (CNS). They are involved in numerous physiological processes, such as movement, respiration, and processing of sensory information, as well as in regulation of neuronal excitability in different brain regions. GlyRs play important role in the maintenance of excitatory/inhibitory balance in the hippocampus and participate in the development of various brain pathologies. In the present study, we have examined a surface expression of GlyRs by pyramidal neurons and astrocytes in control and after 30 min of oxygen-glucose deprivation (OGD) in the organotypic culture of hippocampal slices. Our investigation has demonstrated a decrease in GlyR-positive staining associated with pyramidal neurons and relative stability of GlyRs expression at the surface of astrocytes 4 hs after OGD. These data indicate that GlyRs dysfunction may represent a significant additional factor leading to enhanced neuronal damage induced by OGD. Pharmacological modulation of GlyRs is a promising venue of research for the correction of negative consequences of oxygen-glucose deficiency.

The control of ligand-gated receptors with light using photochromic compounds has evolved from the first handcrafted examples to accurate, engineered receptors, whose development is supported by rational design, high-resolution protein structures, comparative pharmacology and molecular biology manipulations. Photoswitchable regulators have been designed and characterized for a large number of ligand-gated receptors in the mammalian nervous system, including nicotinic acetylcholine, glutamate and GABA receptors. They provide a well-equipped toolbox to investigate synaptic and neuronal circuits in all-optical experiments. This focused review discusses the design and properties of these photoswitches, their applications and shortcomings and future perspectives in the field. Linked articles This article is part of a themed section on Nicotinic Acetylcholine Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc.

Glycine receptors (GlyRs) are indispensable to maintain excitatory/inhibitory balance in neuronal circuits controlling reflex and rhythmic motor behaviors. Here we have developed Glyght, the first GlyR ligand controlled with light. It is selective over other cys-loop receptors, active in vivo, and displays an allosteric mechanism of action. The photomanipulation of glycinergic neurotransmission opens new avenues to understand inhibitory circuits in intact animals, and to develop drug-based phototherapies.

Glycine receptors (GlyRs) belong to the family of ligand-gated cys-loop receptors and effectuate fast inhibitory neurotransmission in central nervous system (CNS). They are involved in numerous physiological processes, such as movement, respiration, and processing of sensory information, as well as in regulation of neuronal excitability in different brain regions. GlyRs play important role in the maintenance of excitatory/inhibitory balance in the hippocampus and participate in the development of various brain pathologies. In the present study, we have examined a surface expression of GlyRs by pyramidal neurons and astrocytes in control and after 30 min of oxygen-glucose deprivation (OGD) in the organotypic culture of hippocampal slices. Our investigation has demonstrated a decrease in GlyR-positive staining associated with pyramidal neurons and relative stability of GlyRs expression at the surface of astrocytes 4 hs after OGD. These data indicate that GlyRs dysfunction may represent a significant additional factor leading to enhanced neuronal damage induced by OGD. Pharmacological modulation of GlyRs is a promising venue of research for the correction of negative consequences of oxygen-glucose deficiency.

For years, our ability to study pathological changes in neurological diseases has been hampered by the lack of relevant models until the recent groundbreaking work from Yamanaka's group showing that it is feasible to generate induced pluripotent stem cells (iPSCs) from human somatic cells and to redirect the fate of these iPSCs into differentiated cells. In particular, much interest has focused on the ability to differentiate human iPSCs into neuronal progenitors and functional neurons for relevance to a large number of pathologies including mental retardation and behavioral or degenerative syndromes. Current differentiation protocols are time-consuming and generate limited amounts of cells, hindering use on a large scale. We describe a feeder-free method relying on the use of a chemically defined medium that overcomes the need for embryoid body formation and neuronal rosette isolation for neuronal precursors and terminally differentiated neuron production. Four days after induction, expression of markers of the neurectoderm lineage is detectable. Between 4 and 7 days, neuronal precursors can be expanded, frozen, and thawed without loss of proliferation and differentiation capacities or further differentiated. Terminal differentiation into the different subtypes of mature neurons found in the human brain were observed. At 6–35 days after induction, cells express typical voltage-gated and ionotrophic receptors for GABA, glycine, and acetylcholine. This specific and efficient single-step strategy in a chemically defined medium allows the production of mature neurons in 20–40 days with multiple applications, especially for modeling human pathologies.

Ginkgo biloba extract is a neuroactive agent that is widely used for correction of age-associated impairment of memory, attention deficit and other cognitive functions. It has been shown that ginkgolides and bilobalides, Ginkgo biloba extract components, are potent blockers of glycine receptors (Kondratskaya et al., 2002, Hawthorne et al., 2006). However, the effect of ginkgolic acid, the other Ginkgo biloba extract constituent, on ligand-gated ion channels was not investigated. In the present study, using patch-clamp technique and transient transfection of different subunits in CHO cells we have shown that glycine receptors (GlyRs) are modulated by ginkgolic acid in a subunit-specific manner. After pre-application of ginkgolic acid (0.5-2 min), glycine-induced currents mediated by α1 GlyRs were strongly potentiated, while currents mediated by α2 GlyRs exhibited weak inhibition. There was no significant effect of ginkgolic acid on amplitudes of currents mediated by α3 GlyRs or on GABAARs composed of 1α/1β/2γ subunits. In order to further investigate subunit-specific effect of ginkgolic acid we have focused on possible interaction sites for this compound inside different GlyR domains. We found that mutation of three residues (T59A/A261G/A303S) in α2 subunit can convert the inhibitory action of ginkgolic acid into potentiation. Indeed, application of ginkgolic acid to cells expressing α2 T59A/A261G/A303S subunits resulted in an increase of responses to low concentrations of glycine and abolishment of the inhibitory effect, typical for wild type α2 GlyR. Our results suggest that (i) ginkgolic acid selectively enhances the function of α1 GlyRs and attenuates the function of α2 GlyRs, (ii) mutation of α2 subunit converts effect of ginkgolic acid from inhibition to potentiation.

Post-ischemic injury of the hippocampus unrolls at different levels and has both functional and structural implications. The deficiency in neuron energy metabolism is an initiating factor. We performed transmission electron microscopic (TEM) comparative analysis of mitochondria in excitatory spine synapses in CA1 stratum radiatum and CA3 hippocampal areas after 5 min of global cerebral ischemia in Mongolian gerbils, 4 and 7 days after reperfusion. Electron microscopy and unbiased morphometric methods were used to evaluate synaptic plasticity, and the number and size of mitochondria in synaptic terminals. We compared the morphological organization of mitochondria in presynaptic terminals between CA1 and CA3 areas in control and post-ischemic condition according to the following morphometric parameters: mitochondrial volume fraction, mitochondrial frequency in CA1 and CA3 terminals, mean number of mitochondria per presynaptic terminal, frequency of damaged mitochondria in terminals, and density of presynaptic terminals. Our ultrastructural study revealed statistically significant differences in morphometric parameters between CA1 and CA3 areas in control conditions, as well as in post-ischemic conditions. Also, we found temporal differences in measured parameters obtained 4 and 7 days after reperfusion. This study showed significant morphological differences in the organization of mitochondria in excitatory spine synapses between CA1 and CA3 areas, which corresponded with already known differences in functionality and sensitivity to the ischemic insult. Our conclusion is that revealed post-ischemic changes in mitochondrial distribution in presynaptic CA1 and CA3 terminals could be an indicator of hippocampal metabolic dysfunction and synaptic plasticity. Anat Rec,, 2011. © 2011 Wiley-Liss, Inc.

Optogenetic and photopharmacological tools to manipulate neuronal inhibition have limited efficacy and reversibility. We report the design, synthesis, and biological evaluation of Fulgazepam, a fulgimide derivative of benzodiazepine that behaves as a pure potentiator of ionotropic γ-aminobutyric acid receptors (GABA A Rs) and displays full and reversible photoswitching in vitro and in vivo. The compound enables high-resolution studies of GABAergic neurotransmission, and phototherapies based on localized, acute, and reversible neuroinhibition.