M-741 (3,15 di-[5,5-dimethyl-3-N[-(cyclopropylmethylinium)-(N-propylinium]-1-cyclohex-1-enyl]-7,11,18,21-tetraoxa-3,15-diazatrispiro [5.2.2.5.2.2] heneicosane produces hepatotoxicity in rats following intravenous administration. Hepatocellular pathology is characterized by parenchymal cell necrosis and inflammatory cell infiltration. Electron microscopic evaluation could not identify any treatment-related effects on mitochondria or the production of cytoplasmic lysosomal lamellar bodies. The M-741-induced hepatotoxicity is not modified by manipulations of nutritional status (fasting), hepatic enzyme induction (phenobarbital) or interference (glutathione depletion) with potential detoxication pathways. The M-741 pharmacokinetic profile is best described by a three compartment model and displays a rapid distribution and terminal elimination. In contrast, hepatic tissue concentrations of M-741 are elevated following administration and prolonged tissue residence is observed. These data are consistent with rapid hepatic uptake and bioaccumulation of M-741. The M-741 hepatotoxicity can be modeled in precision-cut hepatic slices in dynamic culture at concentrations which are measured during in vivo toxication. The toxicophore of the M-741 is the enamino-ether quat moiety and not the spiro-diamine portion of the molecule. Structural analogs of the enamino-ether quat also produced in vitro hepatotoxicity. The in vitro toxicity of M-741 demonstrated temperature dependence and the toxicity could be initiated by short, 30 to 60 minute, pulsed exposure of the hepatic slices to M-741. These findings are consistent with rapid hepatocellular transport of M-741. Hepatic slices accumulated intracellular levels of M-741 during pulsed exposure. M-741 was transported against a concentration gradient and the transport displayed temperature dependence. Known substrates for cationic transport in hepatocytes, d-tubocurarine and triethylme thylammonium bromide, did not reduce M-741 uptake in hepatic slice competition experiments, however, the sensitivity of these measurements may have been inadequate to determine competitive effects on initial uptake velocities. Alternatively, M-741 may be transported intracellularly by absorptive endocytosis as has been demonstrated for other cationic compounds.

The need to develop selective compounds for 5-HT receptors requires further elucidation of structure-activity relationships involved with receptor recognition and activation. In particular, there is great interest in the involvement of the 5-HT₁(A) receptor in therapeutic treatment of conditions such as anxiety, depression and hypertension. Despite previous attempts to define the pharmacophore for 5-HT₁(A) receptors, the structural requirements for 5-HT₁(A) receptor activity remain largely unexplored. In fact, no selective 5-HT₁(A) antagonists currently exist. Binding assays provide important information concerning receptor affinity, but do not reflect functional consequences of receptor interaction. One of the functional correlates of the 5-HT₁(A) receptor is the inhibition of forskolin-stimulated adenylate cyclase (FSC). The objective of this dissertation was to assess the merits of using the FSC assay as a functional screen to determine 5-HT₁(A) intrinsic activity of novel compounds. From the variety of compounds tested, it was apparent that the FSC assay can be used as an in vitro functional screen for assessing 5-HT₁(A) agonistic and antagonistic, as well as partial agonistic, activity. All putative 5-HT₁(A) agonists tested in the FSC assay had potencies (EC₅₀) that were less than their respective 5-HT₁(A) binding affinities (Kᵢ). The use of pindolol to block 5-HT₁(A) receptors, and therefore cause a rightward shift in any observed inhibition curve, provided qualitative evidence of 5-HT₁(A) interaction. Quantitative evaluation of 5-HT₁(A) activity had to be approached cautiously, due to possible complications arising from non-5-HT₁(A)-mediated inhibition. The data for a series of enanatiomers of 8-OH-DPAT and its analogs supported a recently proposed model for the 5-HT₁(A) agonist pharmacophore. Also, within the enantiomeric pairs exhibiting stereoselectivity, the compound with the higher 5-HT₁(A) affinity also possessed greater 5-HT₁(A) agonistic activity. Within a series of tetrahydropyridylindoles (THPI), the 4-THPI analogs generally had both greatest 5-HT₁(A) affinity and agonistic activity, although there were exceptions to this trend. Several compounds, whose structures were based on the small molecules of the 5-HT₁(A) agonist 8-hydroxy-2-(di-n-propylaminotetralin) (8-OH-DPAT) and 5-HT itself, demonstrated potential antagonistic activity in the FSC assay. Further analysis of an extended series of these compounds in the FSC assay is required to establish specific conclusions concerning the 5-HT₁(A) pharmacophore.

This research evaluated the effect of air velocity and other cure parameters, such as temperature and oven air recirculation, on the release of 4-Phenylcyclohexene from carpet samples coated with a composite XSBR latex. This kind of carpet is similar to that used in industries. The effect of latex frothing process on the reduction of 4-PCH from SBR latex samples was also investigated. It has been found that both air velocity and oven temperature were important cure parameters in 4-PCH reduction. Air velocity was the dominating factor in the removal of 4-PCH from carpet back-coated with latex composite. At oven temperature of 225°F, the content of 4-PCH of carpet samples can be reduced by 80% when it was exposed on an air velocity of 200 fpm. While the 4-PCH content can only reduced by 15% if it is not exposed to flowing air. Frothing process also had significant influence on the release of 4-PCH from XSBR latex.

In the present study, we have demonstrated that PBR sites are present in the microsomal fraction from rat liver, heart and kidney, which we have named the non-mitochondrial PBR. The non-mitochondrial PBR was pharmacologically characterized by porphyrin competition experiments. The ability of porphyrins to differentially compete for specific (3H) Ro5-4864 or (3H) PK11195 binding demonstrated that the mitochondrial and non-mitochondrial PBR have different affinities for selected porphyrin compounds. These results suggest that the mitochondrial and non-mitochondrial PBR have a different binding pharmacology and support the existence of a non-mitochondrial PBR site. On the other hand, porphyrins show different potencies in competing for specific (3H) Ro5-4864 and (3H) PK11195 binding in the same fraction. In addition, the results of saturation experiments show that there are more PK11195 binding sites than Ro5-4864 binding sites in all fractions examined. This observation suggests that the sites labeled by (3H) Ro5-4864 and (3H) PK11195 are not identical.

Hepatic drug clearance (HDC) plays a fundamental role in the metabolism and disposition of xenobiotics. Numerous methods of assessing HDC have been proposed in an effort to elucidate the genetic and extra-genetic effects upon xenobiotic biotransformation. One such method incorporates the co-administration of three model compounds to assess three unique parameters of HDC; namely, hepatic blood flow, Phase I (microsomal-mediated oxidative) metabolism, and Phase II (synthesis/conjugation) reactions. Pharmacokinetic studies of indocyanine green, antipyrine, and lorazepam were undertaken to assess each HDC parameter, respectively. This method was utilized to define differences in HDC between normal, healthy, young males and an older group with cancer.

The major objective of this work was development of procedures for the quantification of cutaneous microsomal cytochrome P-450 mediated metabolism using model substrates. The Fischer-344 rat was the species of choice for this developmental work. The versatility of this method was demonstrated through its application to a variety of species including man. An extension of the rat microsomal work entailed the use of inducing agents, specifically, the Aroclor series of polychlorinated biphenyls. The effects of these inducing agents were demonstrated and characterized in skin and liver with respect to microsomal marker enzymes and substrate metabolism. In addition to the microsomal studies, the use of rat skin explants in culture, provided a simple system for the characterization of phase I and phase II coupled metabolism of the model substrates.

New radioligand [4'-¹²⁵I-[Phe⁴]DPDPE ([¹²⁵I] DPDPE) was prepared from [4'-NH₂-Phe⁴] DPDPE. Its binding characteristics were determined using rat brain membranes. Saturation studies showed one binding site with K(d) = 421 ± 67 pM and B(max) = 36.4 ± 2.7 fmol/mg protein. Kinetic studies showed association rate constant k₊₁ = 5.80 ± 0.88 x 10⁷ M⁻¹ min⁻¹ and dissociation rate constant k-1 = 0.917 ± 0.117 x 10⁻² min⁻¹. Competitive inhibition studies showed high selectivity for delta opioid receptors. Distribution of [¹²⁵I] DPDPE labeled sites in rat brain (autoradiography) is consistent with previous results using other radioligands. The inhibition constants of [¹²⁵I] DPDPE binding by delta selective ligands were similar in mouse brain and mouse vas deferens, and no significantly different values were determined in rat brain. These data indicate that [¹²⁵I] DPDPE due to high specific activity is valuable for characterization of delta opioid receptors.

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