Cutting frozen sections of large (greater than 60 cc) blocks of monkey brain using the conventional procedures of infiltration with 30% sucrose as a cryoprotectant before freezing with pulverized dry ice often produces unacceptable levels of freezing artifact (FA) caused by displacement of tissue by ice crystals. Experiments investigating FA utilized perfusion-fixed brains from 46 monkeys and spanned combinations of cryoprotectants (glycerol, sucrose), freezing methods (dry ice or -75 degrees C isopentane), and fixatives (10% formalin, Karnovsky's or Timm's). The effects were evaluated by rating of FA severity in frozen sections of whole monkey brains. Minor FA appears as enlarged capillaries, more serious FA as large vacuoles, and both first appear midway between the periphery and center of the block. Stronger fixatives increased the severity of freezing artifact. The best method for eliminating FA was graded infiltration with up to 20% glycerol and 2% DMSO (in buffer or fixative), followed by rapid freezing in -75 degrees C isopentane. Although using a glycerol-DMSO infiltration before conventional freezing with pulverized dry ice or using conventional sucrose infiltration before freezing in isopentane gave better results than sucrose infiltration and dry-ice freezing, only the combination of glycerol-DMSO infiltration and freezing in isopentane produced consistently excellent results and virtually eliminated freezing artifact. To determine the effect of freezing with dry ice or isopentane on the rate of cooling in large blocks of CNS tissue, thermocouples were embedded in an 80-cc block of albumin-gelatin and frozen with the two methods. The rate of cooling (-3.5 degrees C/min) was twice as fast using isopentane.
These data suggest nosocomial transmission of multidrug-resistant tuberculosis occurred from patient to patient and from patient to health care worker and underscore the need for effective acid-fast bacilli isolation facilities and adherence to published infection control guidelines in health care institutions.
The centrifugal projections from the various subdivisions of the anterior olfactory nucleus (AON) can be categorized into four groups based on the organization of terminal fields in the main olfactory bulb (MOB). Pars lateralis and dorsalis have bilaterally asymmetric laminar projections to the MOB. The ipsilateral projections terminate primarily in the superficial half of the granule cell layer and in the deep third of the glomerular layer, whereas the contralateral projections terminate primarily in the superficial half of the granule cell layer and do not extend into the glomerular layer. Pars ventralis and posterior have bilaterally symmetric laminar projections with heavy terminations both in the superficial half of the granule cell layer and in the deep third of the glomerular layer. Pars medialis sends predominantly ipsilateral projections to the deep half of the granule cell layer. Pars externa has predominantly contralateral projections with a very narrow terminal field immediately deep to the internal plexiform layer. The projections to the MOB from the ventral hippocampal rudiment (HR) and the piriform cortex (PC) are exclusively ipsilateral. The projections from the ventral HR terminate primarily in the deep half of the granule cell layer. The projections from the PC also terminate predominantly in the granule cell layer, but there is a progressive shifting of terminal fields from the superficial half of this layer toward deeper regions for centrifugal axons arising from progressively more caudal levels of the PC. The laminar termination patterns of cortical afferents to the ipsilateral MOB thus are correlated with the mediolateral axis of the olfactory peduncle and the rostrocaudal axis of the piriform cortex. The centrifugal axons from these various sources enter directly into the granule cell layer of the caudal MOB or pass through the internal plexiform layer of the accessory olfactory bulb to reach the middle and anterior part of the MOB. We have termed these two routes the final common bulb pathway. The centrifugal axons from the laterally situated sources join the anterior and bulbar limbs of the anterior commissure before entering the final common bulbar pathway. In contrast, the centrifugal axons from pars medialis and the ventral HR travel diffusely in the cellular layer of the ipsilateral olfactory peduncle. A small component of the centrifugal projections from the PC travels in association with the lateral olfactory tract.
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