Since Dale and Laidlaw (1) first reported the contraction of smooth muscle by histamine, the pharmacological significance of this phenomenon has been extensively investigated. Three subtypes of histamine receptor (H1, H2, and H3) are known. The H1 receptor was identified by Ash and Schild (2) and H1 receptor antagonists have been used in the therapy of many allergic diseases, including urticaria, allergic rhinitis, pollenosis, and bronchial asthma. In peripheral tissues, the histamine H1 receptor mediates the contraction of smooth muscles, increase in capillary permeability due to contraction of terminal venules, and catecholamine release from adrenal medulla (3), as well as mediating neurotransmission in the central nervous system (4). Although signal transduction of the H1 receptor through Ca2l mobilization via an increase in the intracellular inositol 1,4,5-trisphosphate level has been extensively investigated (5,6), little is known about the molecular structure of the histamine H1 receptor. Recently, Isolation ofPoly(A)+ RNA. Total RNA was extracted by the acid guanidinium isothiocyanate/phenol/chloroform method (11). Poly(A)+ RNA was isolated by chromatography on oligo(dT)-cellulose (12).Expression Cloning of Histamine HI Receptor cDNA. Bovine adrenal medullary poly(A)+ RNA ('180 ug) was sizefractionated on a 5-25% (wt/vol) sucrose-density gradient.An aliquot (1 p1L) of each poly(A)+ RNA fraction (20,ul) was injected into Xenopus oocytes, and electrophysiological assay by measuring Ca2+-dependent inward Cl-currents was done as described (9). The fraction that showed the highest histamine-induced inward Cl-currents was used for oligo(dT)-primed cDNA synthesis. Double-stranded cDNAs of >2-kilobase (kb) pairs were size-selected by agarose gel electrophoresis followed by elution with Geneclean II (Bio 101, La Jolla, CA) and were ligated into AZAPII (Stratagene) at the EcoRI site. The library was divided and amplified in 65 pools of "'-20,000 independent clones each. In vitro transcription was done essentially according to the procedure ofJulius et al. (13). RNA transcripts (=5 ng) from each pool were individually injected into Xenopus oocytes. After incubation for 1-2 days, the oocytes were tested for inward Cl-currents induced by 100 ,uM histamine under a voltage clamp at -60 mV. The single positive pool of 20,000 clones was progressively subdivided into smaller pools of 8000, 4000, 400, and 15 clones until finally a single clone was obtained. cDNA encoding the histamine H1 receptor was sequenced by the M13 chain-termination method (14) using a DNA sequencer (model 370A, Applied Biosystems). The sequence homology search was done by using DNASIS (Hitachi Software Engineering, Yokohama, Japan).

A nonnucleoside reverse transcriptase (RT) inhibitor, GW420867, was tested for postexposure prophylaxis (PEP) in rhesus macaques experimentally infected with 100 50% tissue culture infective doses of a chimeric simian/human immunodeficiency virus (SHIV) containing the RT gene of HIV-1 (SHIV-RT). Animals were either mock treated, or treated for 4 weeks starting at 8 or 24 h postinfection (p.i.) with GW420867. While such therapy led to undetectable plasma viremia in three of six monkeys, a transient plasma viremia was noted in the other three treated animals at 2 to 4 weeks following cessation of therapy. Following this transient viremia all drug-treated animals showed low or undetectable levels of plasma viremia up to the last sample examined at 90 weeks p.i. Despite low and/or undetectable viremia, virus-specific cytotoxic T lymphocyte and viral Envspecific proliferative responses were seen in the peripheral blood mononuclear cells of both mock-and drugtreated animals as early as 3 weeks p.i. Such virus-specific cellular responses, however, were better maintained in the drug-treated animals than the mock-treated animals. In contrast to the virus-specific cellular response, the magnitude and kinetics of virus specific humoral responses appeared to correlate with the detection of viremia. These data support the view that a short-term PEP with GW420867 permits the generation and maintenance of long-lasting virus-specific cell-mediated immune responses while markedly reducing viral loads to undetectable levels for a prolonged period of time (90 weeks) and leads to long-term disease protection. This model provides a unique means to define mechanisms and correlates of disease protection.

No abstract

No abstract

No abstract