Allyn C Howlett

• Wake Forest University

Cannabinoid Receptor Interacting Protein isoform 1b (CRIP1b) is known to interact with the CB1 receptor. Alternative splicing of the CNRIP1 gene produces CRIP1a and CRIP1b with a difference in the third exon only. Exons 1 and 2 encode for a functional domain in both proteins. CRIP1a is involved in regulating CB1 receptor internalization, but the fu...

G protein-coupled receptors (GPCRs) are important regulators of cellular signaling functions and therefore are a major target for drug discovery. The CB1 cannabinoid receptor is among the most highly expressed GPCRs in neurons, where it regulates many differentiated neuronal functions. One model system for studying the biochemistry of neuronal resp...

The CB1 and CB2 cannabinoid receptors (CB1R, CB2R) are members of the G protein-coupled receptor (GPCR) family that were identified over 20 years ago. CB1Rs and CB2Rs mediate the effects of Δ⁹-tetrahydrocannabinol (Δ⁹-THC), the principal psychoactive constituent of marijuana, and subsequently identified endogenous cannabinoids (endocannabinoids) an...

Recent studies have used conditional knockout mice to selectively delete the D2 autoreceptor; however, these approaches result in global deletion of D2 autoreceptors early in development. The present study takes a different approach using RNA interference (RNAi) to knockdown the expression of the D2 receptors (D2R) in the substantia nigra (SN), inc...

Cannabinoid Receptor Interacting Protein1a (CRIP1a) is a CB1 receptor (CB1R) distal C-terminus-associated protein that alters CB1R interactions with G-proteins (Blume et al.,2015; Smith et al.,2015). We tested the hypothesis that CRIP1a is capable of also altering CB1R interactions with β-arrestin proteins that interact with the CB1R at the C-termi...

Dysregulated expression and function of dopamine D2 receptors (D2Rs) are implicated in drug addiction, Parkinson's disease and schizophrenia. In the current study, we examined whether D2Rs are modulated by regulator of G protein signaling 2 (RGS2), a member of the RGS family that regulates G protein signaling via acceleration of GTPase activity. Us...

Cannabinoid receptor interacting protein 1a ( CRIP 1a) is a CB 1 receptor ( CB 1 R) distal C‐terminus‐associated protein that modulates CB 1 R signaling via G proteins, and CB 1 R down‐regulation but not desensitization (Blume et al . [2015] Cell Signal ., 27, 716–726; Smith et al . [2015] Mol. Pharmacol ., 87, 747–765). In this study, we determine...

CB1 cannabinoid receptors are expressed on vagal afferent fibers and neurons within the solitary tract nucleus (NTS), providing anatomical evidence for their role in arterial baroreflex modulation. To better understand the relationship between the brain renin-angiotensin system (RAS) and endocannabinoid expression within the NTS, we measured dorsal...

CB1 cannabinoid receptors (CB1R) are found in abundance in the neurons where they can affect neuronal plasticity by modulating intracellular signal transduction mechanisms including cAMP (1) and Ca2+ (2,3) dynamics. The presence of the G-protein coupled calcium-sensing receptor (CaS) has been previously demonstrated in N18TG2 neuroblastoma cells (4...

To improve overall healthcare and to reduce health disparities, efforts must focus on increasing the diversity of personnel trained in the biomedical sciences. Here, we describe the development, implementation, and relative outcomes of three pipeline training programs in biomedical sciences research designed to increase workforce diversity institut...

Background: CB1 cannabinoid receptors (CB1Rs) stimulate Gi/o-dependent signaling pathways. CB1R-mediated cAMP increases were proposed to result from Gs activation, but CB1R-stimulated GTPγS binding to Gs has not heretofore been investigated. Methods: Three models of CB1R-stimulated cAMP production were tested: pertussis toxin disruption of Gi/o...

Hypertension resulting from elevated brain angiotensin (Ang) II is associated with impaired functioning of neural reflexes regulating sympathetic and parasympathetic outflow. Restoration of normal baroreflex sensitivity (BRS) for control of heart rate (HR) is achieved in a rat model of Ang II - dependent hypertension [(mRen2)27 transgenic rats] by...

Low-frequency ultrasound (LFU) in the frequency range 30-40 kHz administered using different delivery methods (waterbath and noncontact spray) has shown positive effects on chronic wound healing rates in humans, but the underlying mechanisms are only beginning to be explored. To examine the effects of LFU delivered at 35 kHz on the morphology and m...

The GABAB receptor is a therapeutic target for CNS and neuropathic disorders; however, few preclinical studies have explored effects of chronic stimulation. This study evaluated acute and chronic baclofen treatments on GABAB-activated G-proteins and signaling protein phosphorylation as indicators of GABAB signaling capacity. Brain sections from rat...

Cannabinoid CB1 receptors (CB1R) mediate the presynaptic effects of endocannabinoids in the central nervous system (CNS) and most behavioral effects of exogenous cannabinoids. Cannabinoid Receptor-Interacting Protein 1a (CRIP1a) binds to the CB1R C-terminus and can attenuate constitutive CB1R-mediated inhibition of Ca(2+) channel activity. We now d...

As they age, Sprague-Dawley (SD) rats develop elevated systolic blood pressure associated with impaired baroreflex sensitivity (BRS) for control of heart rate. We previously demonstrated in young hypertensive (mRen2)27 rats that impaired BRS is restored by CB1 cannabinoid receptor blockade in the solitary tract nucleus (NTS), consistent with elevat...

CB1 cannabinoid receptors (CB1R) are one of the most abundantly expressed G protein coupled receptors (GPCR) in the CNS and regulate diverse neuronal functions. The identification of GPCR interacting proteins has provided additional insight into the fine-tuning and regulation of numerous GPCRs. The cannabinoid receptor interacting protein 1a (CRIP1...

Objectives To investigate the role of COX-derived prostanoids and the endocannabinoids 2-arachidonoylglycerol (2-AG) and anandamide (AEA) on uterine vascular control during early pregnancy in a rodent model of preeclampsia. Methods The transgenic female rat containing the human angiotensinogen (hAGN) gene mated with the male transgenic containing h...

Isolation and structure elucidation of most of the major cannabinoid constituents - including Δ(9)-tetrahydrocannabinol (Δ(9)-THC), which is the principal psychoactive molecule in Cannabis sativa - was achieved in the 1960s and 1970s. It was followed by the identification of two cannabinoid receptors in the 1980s and the early 1990s and by the iden...

We investigated acute and chronic effects of CB1 cannabinoid receptor blockade in renin-angiotensin system-dependent hypertension using rimonabant (SR141716A), an orally active antagonist with central and peripheral actions. In transgenic (mRen2)27 rats, a model of angiotensin II-dependent hypertension with increased body mass and insulin resistanc...

The progression from Raphael Mechoulam's identification of Δ9-tetrahydrocannabinol as the primary psychoactive component of cannabis, through the design of cannabinoid analogues by academic and pharmaceutical chemists, to the discovery of CB1 and CB2 cannabinoid receptors, brings us to modern pharmacological investigation. The CB1 receptor interact...

Increased vascular sensitivity to angiotensin II (Ang II) is a marker of a hypertensive human pregnancy. Recent evidence of interactions between the renin-angiotensin system and the endocannabinoid system suggests that anandamide and 2-arachidonoylglycerol may modulate Ang II contraction. We hypothesized that these interactions may contribute to th...

Skin fibroblasts comprise the first barrier of defense against wounds, and tobacco products directly contact the oral cavity. Cultured human dermal fibroblasts were exposed to smokeless tobacco extract (STE), total particulate matter (TPM) from tobacco smoke, or nicotine at concentrations comparable to those found in these extracts for 1 h or 5 h....

The contribution of individual CB receptors (CB1 R and CB2 R) to normal micturition has not been clearly defined. Our goal was to study if differences in urodynamic parameters or in vitro bladder contractility can be demonstrated between CB2 R knockout (CB2 RKO) and C57BL/6J control (wild type, WT) mice. Female WT and CB2 RKO mice underwent bladder...

Dopamine D2 autoreceptors located on the midbrain dopaminergic neurons modulate dopamine (DA) neuron firing, DA release, and DA synthesis through a negative feedback mechanism. Dysfunctional D2 autoreceptors following repeated drug exposure could lead to aberrant DA activity in the ventral tegmental area (VTA) and projection areas such as nucleus a...

The serine hydrolase α/β hydrolase domain 6 (ABHD6) has recently been implicated as a key lipase for the endocannabinoid 2-arachidonylglycerol (2-AG) in the brain. However, the biochemical and physiological function for ABHD6 outside of the central nervous system has not been established. To address this, we utilized targeted antisense oligonucleot...

The effect of the endocannabinoid system (ECS) on weight and energy metabolism is well accepted, but long term contributions of the ECS to cardiovascular regulation are not established. Evidence for receptor and signaling interactions between the ECS and the renin-angiotensin system (RAS) in brain and vasculature is emerging. In transgenic (mRen2)2...

Complete artificial saliva (CAS) is a saliva substitute often used as a vehicle for test articles, including smokeless tobacco products. In the course of a study employing normal adult human dermal fibroblasts (HDFa) as a model in vitro, we discovered that CAS as a vehicle introduced a significant change in the expression of proinflammatory cytokin...

Tyrosine phosphorylation (Tyr-P) of focal adhesion kinase (FAK) regulates FAK activation. Phosphorylated FAK Tyr 397 binds Src family kinases (Src), which in turn directly phosphorylate FAK Tyr 576/577 to produce maximal FAK enzymatic activity. CB1 cannabinoid receptors (CB1) are abundantly expressed in the nervous system and influence FAK activati...

Normal aging may limit the signaling efficacy of certain GPCRs by disturbing the function of specific Gα-subunits and leading to deficient modulation of intracellular functions that subserve synaptic plasticity, learning and memory. Evidence suggests that Gα(q/11) is more sensitive to the effects of aging relative to other Gα-subunits, including Gα...

Although biochemical and physiological evidence suggests a strong interaction between striatal CB(1) cannabinoid (CB(1) R) and D(2) dopamine (D(2) R) receptors, the mechanisms are poorly understood. We targeted medium spiny neurons (MSNs) of the indirect pathway using shRNA to knockdown either CB(1) R or D(2) R. Chronic reduction in either receptor...

Lower extremity amputations and foot ulcers are complications associated with diabetes, and have been shown to affect diabetic African Americans (AA) three times as often as diabetic non-Hispanic Whites (NHW). Possible causes for the increased risk include ethnic differences in structure and function within the dermis of the lower extremity. Testin...

BACKGROUND AND PURPOSESignalling networks that regulate the progression of cannabinoid CB1 receptor‐mediated extracellular signal‐regulated kinase (ERK) activation in neurons are poorly understood. We investigated the cellular mechanisms involved in CB1 receptor‐stimulated ERK phosphorylation in a neuronal cell model. EXPERIMENTAL APPROACHMurine N1...

The CB(1) cannabinoid receptor is regulated by its association with membrane microdomains such as lipid rafts. Here, we investigated the role of palmitoylation of the CB(1) receptor by analysing the functional consequences of site-specific mutation of Cys(415) , the likely site of palmitoylation at the end of helix 8, in terms of membrane associati...

Background and purpose: Signalling networks that regulate the progression of cannabinoid CB(1) receptor-mediated extracellular signal-regulated kinase (ERK) activation in neurons are poorly understood. We investigated the cellular mechanisms involved in CB(1) receptor-stimulated ERK phosphorylation in a neuronal cell model. Experimental approach:...

This review evaluates the cellular mechanisms of constitutive activity of the cannabinoid (CB) receptors, its reversal by inverse agonists, and discusses the pitfalls and problems in the interpretation of the research data. The notion is presented that endogenously produced anandamide (AEA) and 2‐arachidonoylglycerol (2‐AG) serve as autocrine or pa...

There are at least two types of cannabinoid receptors (CB(1) and CB(2)). Ligands activating these G protein-coupled receptors (GPCRs) include the phytocannabinoid Δ(9)-tetrahydrocannabinol, numerous synthetic compounds, and endogenous compounds known as endocannabinoids. Cannabinoid receptor antagonists have also been developed. Some of these ligan...

Previous research has demonstrated that concurrent systemic administration of CB(1) cannabinoid and mu-opioid receptor agonists increases feeding in rats. However, the possible neural loci of this cooperative effect have yet to be identified. These studies tested whether the nucleus accumbens shell may be one site of the interactive effects of opio...

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

CB1 receptors are G-protein coupled receptors (GPCRs) abundant in neurons, in which they modulate neurotransmission. The CB(1) receptor influence on memory and learning is well recognized, and disease states associated with CB(1) receptors are observed in addiction disorders, motor dysfunction, schizophrenia, and in bipolar, depression, and anxiety...

The endocannabinoids anandamide and 2-arachidonoylglycerol are lipid mediators that signal via CB(1) and CB(2) cannabinoid receptors and Gi/o-proteins to inhibit adenylyl cyclase and stimulate mitogen-activated protein kinase. In the brain, CB(1) receptors interact with opioid receptors in close proximity, and these receptors may share G-proteins a...

The proximal portion of the C-terminus of the CB 1 cannabinoid receptor is a primary determinant for G-protein activation. A 17 residue proximal C-terminal peptide (rodent CB1 401–417), the intracellular loop 4 (IL4) peptide, mimicked the receptor's G-protein activation domain. Because of the importance of the cationic amino acids to G-protein acti...

In our previous studies, CB1 cannabinoid receptor agonists stimulated production of cyclic GMP and translocation of nitric oxide (NO)-sensitive guanylyl cyclase in neuronal cells (Jones et al., Neuropharmacology 54:23–30, 2008). The purpose of these studies was to elucidate the signal transduction of cannabinoid-mediated neuronal nitric oxide synth...

CB1 and CB2 cannabinoid receptors are associated with Gi/o proteins to activate signal transduction pathways that include inhibition of adenylyl cyclase, activation of mitogen activated protein kinase (MAPK), and regulation of ion channels (CB1 only). Agonists for these receptors include structurally diverse cannabinoid, aminoalkylindole, and eicos...

The CB1 cannabinoid receptor is a G-protein-coupled receptor (GPCR) that can be stimulated by cannabinoid and nonclassical cannabinoids, aminoalkylindoles, and endocannabinoids. Constitutive activity of CB1 receptors allows inverse agonists such as arylpyrazoles to modulate activity in the absence of an agonist, and can allow sequestration of Gi/o...

Endocannabinoid, opioid, and dopamine systems interact to exhibit cannabinoid receptor neuromodulation of opioid peptides and D(4) dopamine receptor gene expression in CB(1)-cannabinoid-deficient mouse striatum. Using CB(1)-transgenic mice, we examine primary age-sex influences and interactions on opioid and dopamine system members' gene expression...

Compound 6 poses within the CB1 receptor having optimal Affinity lowest energy values.

Compound 1 poses within the CB1 receptor having optimal Affinity lowest energy values.

Compound 2 poses within the CB1 receptor having optimal Affinity lowest energy values.

CB1 cannabinoid receptors are G-protein coupled receptors for endocannabinoids including anandamide and 2-arachidonoylglycerol. Because these arachidonic acid metabolites possess a 20-carbon polyene chain as the alkyl terminal moiety, they are highly flexible with the potential to adopt multiple biologically relevant conformations, particularly tho...

Cannabinoid agonists regulate NO and cyclic AMP production in N18TG2 neuroblastoma cells, leading to the hypothesis that neuronal cyclic GMP production could be regulated by CB(1) cannabinoid receptors. NO (nitric oxide)-sensitive guanylyl cyclase (GC) is a heterodimeric cytosolic protein that mediates the down-stream effects of NO. Genes of protei...

The CB1 cannabinoid receptor is a G-protein coupled receptor that has important physiological roles in synaptic plasticity, analgesia, appetite, and neuroprotection. We report the discovery of two structurally related CB1 cannabinoid receptor interacting proteins (CRIP1a and CRIP1b) that bind to the distal C-terminal tail of CB1. CRIP1a and CRIP1b...

The proximal portion of the C-terminus of the CB(1) cannabinoid receptor is a primary determinant for G-protein activation. A 17 residue proximal C-terminal peptide (rodent CB1 401-417), the intracellular loop 4 (IL4) peptide, mimicked the receptor's G-protein activation domain. Because of the importance of the cationic amino acids to G-protein act...

The intracellular C-terminal helix 8 (H8) of the CB1 cannabinoid receptor deviates from the highly conserved NPXXY(X)5,6F G-protein-coupled receptor motif, possessing a Leu instead of a Phe. We compared the signal transduction capabilities of CB1 with those of an L7.60F mutation and an L7.60I mutation that mimics the CB2 sequence. The two mutant re...

(1) This study investigated the functional genomics of glucocorticoid and opioid receptor stimulation in cellular adaptations using a cultured neuronal cell model. (2) Human SH-SY5Y neuroblastoma cells grown in hormone-depleted serum were treated for 2-days with the glucocorticoid receptor-II agonist dexamethasone (30 nM); the mu-opioid receptor ag...

We have previously shown that the endocannabinoid anandamide and its metabolically stable analog (R)-methanandamide produce vasorelaxation in rabbit aortic ring preparations in an endothelium-dependent manner that could not be mimicked by other CB(1) cannabinoid receptor agonists (Am J Physiol 282: H2046-H2054, 2002). Here, we show that (R)-methana...

The rat dorsal root ganglion (DRG) Ca(2+)-sensing receptor (CaR) was stably expressed in-frame as an enhanced green fluorescent protein (EGFP) fusion protein in human embryonic kidney (HEK)293 cells, and is functionally linked to changes in intracellular Ca(2+) concentration ([Ca(2+)](i)). RT-PCR analysis indicated the presence of the message for t...

Scene 2 of TUNNELS. A woman is suffering from withdrawal in her heroin dependence.

Scene 3 of TUNNELS. A drug dealer describes the marketing and sale of crack cocaine.

Scene 1 of TUNNELS. Parents blame each other for their son's behavior leading to a drug-related crime.

Scene 5 of TUNNELS. A pregnant woman, who needs money for her crack habit, steals from a social services worker.

Scene 4 of TUNNELS. Two homeless men discuss the impact of alcohol abuse on their relationship with family.

Scene 6 of TUNNELS. A man is incarcerated for the accidental death of his partner during a crack cocaine binge.

The substance abuse prevention goal of the theatre production "TUNNELS" was to provide community education on substance abuse to an audience in Durham, NC and surrounding communities. The education effort intended to increase awareness and understanding of the risk and protective factors associated with alcohol and other drug use, and to promote pr...

CB(2) cannabinoid receptors exist in immune cells including macrophages. Affinity-purified antibodies against the CB(2) receptor identified a 45 kDa protein in rat brain, human tonsil and rat and mouse microglia, but not mouse N18TG2 neuroblastoma cells. Intracerebroventricular lipopolysaccharide (LPS) increased immunoreactive CB(2) receptors in br...

Neuropathologies that affect our population include ischemic stroke and neurodegenerative diseases of immune origin, including multiple sclerosis. The endocannabinoid system in the brain, including agonists anandamide (arachidonyl ethanolamide) and 2-arachidonoylglycerol, and the CB1 and CB2 cannabinoid receptors, has been implicated in the pathoph...

Key pharmacophoric elements for the (aminoalkyl)indole (AAI) CB1 cannabinoid receptor agonists are the aminoalkyl moiety, the lipophilic aroyl group, and the heterocyclic indole ring. In the present study, the docking space allowed for (R)-[2,3-dihydro-5-methyl-3-[(4-morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazin-6-yl](1-naphthalenyl)methanone...

Antagonism of the CB(1) cannabinoid receptor (CB(1) receptor) by rimonabant (SR141716) reduces self-administration of alcohol and other drugs of abuse in animal models. These findings suggest that the CB(1) receptor may be a target for genetic differences that modify the salient features of rewarding drugs. In the present study, wild-type (CB(1) (+...

The cannabinoid receptor family currently includes twotypes:CB1, characterized in neuronal cells and brain, and CB2, characterized in immune cells and tissues. CB1 and CB2 receptors are members of the superfamily of seven-transmembrane-spanning (7-TM) receptors, having a protein structure defined by an array of seven membrane-spanning helices with...

in this issue of American Journal of Physiology - Heart and Circulatory Physiology , Pacher et al. ([16][1]) characterized the cardiovascular effects of anandamide (AEA) in mice deficient in fatty acid amide hydrolase (FAAH), an enzyme responsible for metabolizing AEA. There was no difference in

The International Union of Pharmacology (IUPHAR) has established a Nomenclature Committee comprised of sub-committees of experts to evaluate types and subtypes of receptors and ion channels in an effort to establish universally accepted nomenclature [Vanhoutte, P.M., Barnard, E.A., Cosmides, G.J., Humphrey, P.P., Spedding, M., Godfraind, T., 1994....

To understand how structurally distinct ligands regulate CB(1) receptor interactions with Gi1, Gi2, and Gi3, we quantified the Galphai and betagamma proteins that coimmunoprecipitate with the CB(1) receptor from a detergent extract of N18TG2 membranes in the presence of ligands. A mixture of A, R, G(GDP) (or G_), and ARG(GDP) (or ARG_) complexes wa...

The cannabinoid receptor family currently includes two types: CB1, characterized in neuronal cells and brain, and CB2, characterized in immune cells and tissues. CB1 and CB2 receptors are members of the superfamily of seven-transmembrane-spanning (7-TM) receptors, having a protein structure defined by an array of seven membrane-spanning helices wit...

CB 1 receptor cellular signal transduction is dependent on the expression of G proteins to which the receptor couples, the potential for precoupling of particular G proteins to the receptors either by scaffolding mechanisms or colocalization in lipid raft domains, and the effector mechanisms that these transducer molecules regulate. This discourse...

To determine the moiety that behaves as the steric trigger to activate the CB(1) cannabinoid receptor, conformational properties of the nonclassical cannabinoid CP55244, one of the most potent CB(1) receptor agonists, were characterized by conformational analysis, rotational barrier calculations, and molecular dynamics (MD) simulations. It was show...

Δ9-Tetrahydrocannabinol from Cannabis sativa is mimicked by cannabimimetic analogs such as CP55940 and WIN55212-2, and antagonized by rimonabant and SR144528, through G-protein-coupled receptors, CB1 in the brain, and CB2 in the immune system. Eicosanoids anandamide and 2-arachidonoylglycerol are the “endocannabinoid” agonists for these receptors....

Association of cannabimimetic compounds such as cannabinoids, aminoalkylindoles (AAIs), and arachidonylethanolamide (anandamide) with the brain cannabinoid (CB(1)) receptor activates G-proteins and relays signals to regulate neuronal functions. A CB(1) receptor homology model was constructed using the published x-ray crystal structure of bovine rho...

Effects of cannabinoid compounds on neurons are predominantly mediated by the CB(1) cannabinoid receptor. Onset of signaling cascades in response to cannabimimetic drugs is triggered by the interaction of the cannabinoid receptor with G(i/o) proteins. Much work has been done to delineate the cannabinoid agonist-induced downstream signaling events;...

Cannabinoid receptors were named because they have affinity for the agonist delta9-tetrahydrocannabinol (delta9-THC), a ligand found in organic extracts from Cannabis sativa. The two types of cannabinoid receptors, CB1 and CB2. are G protein coupled receptors that are coupled through the Gi/o family of proteins to signal transduction mechanisms tha...

Two types of cannabinoid receptor have been discovered so far, CB(1) (2.1: CBD:1:CB1:), cloned in 1990, and CB(2) (2.1:CBD:2:CB2:), cloned in 1993. Distinction between these receptors is based on differences in their predicted amino acid sequence, signaling mechanisms, tissue distribution, and sensitivity to certain potent agonists and antagonists...

The endogenous cannabinoid anandamide (arachidonylethanolamide) produces vasorelaxation in different vascular beds. In the present study, we found that anandamide and a metabolically stable analog, methanandamide, produced dose-dependent (10 nM-10 microM) vasorelaxation of approximately 80% in a rabbit aortic ring preparation in an endothelium-depe...

Zn2+ is required as either a catalytic or structural component for a large number of enzymes and thus contributes to a variety of important biological processes. We report here that low micromolar concentrations of Zn2+ inhibited hormone- or forskolin-stimulated cAMP production in N18TG2 neuroblastoma cells. Similarly, low concentrations inhibited...

N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (SR141716; 1) is a potent and selective antagonist for the CB1 cannabinoid receptor. Using the AM1 molecular orbital method, conformational analysis of 1 around the pyrazole C3 substituent identified four distinct conformations designated Tg, Ts, Cg, and...