There are at least two basal cell populations in the olfactory epithelium that could give rise to olfactory neurons during development, in the normal adult, and after experimentally induced receptor cell death. These populations have been subdivided as horizontal (HBC) and globose (GBC) basal cells on the basis of morphological criteria and by staining with antibodies against cytokeratin. HBCs are positive for cytokeratin while GBCs are negative. We have studied which cell type is induced to divide during receptor cell regeneration stimulated by olfactory bulbectomy using a combination of immunocytochemistry and autoradiography. By examining which population increases its labeling index with 3H-thymidine (3H- TdR) at various times after bulbectomy, it is shown that there is an increase in 3H-TdR uptake in the cytokeratin-negative GBCs with no change in the cytokeratin-positive HBCs. This suggests that the GBCs are specifically induced to divide in response to cues that accompany receptor cell death, and it is thus concluded that these cells are among the precursors of new olfactory receptor neurons.
When rats are treated daily with reserpine or maintained at 4 degrees C, the level of a specific RNA coding for tyrosine hydroxylase is elevated in the adrenal gland. The increase in this specific RNA temporally precedes and is quantitatively equal to the increase in adrenal tyrosine hydroxylase enzyme activity elicited by these treatments. These results suggest that prolonged stress may lead to changes in the levels of specific RNA species in the adrenal gland.
We thank Mr. Walter Dent for photography and Ms. Barbara D'Angelo for preparation of the manuscript. We thank Dr. Charles A. Greer for many helpful and stimulating discussions and for reading of the manuscriot.
CATH.a is a central nervous system (CNS) catecholaminergic cell line derived from a transgenic mouse carrying the SV40 T antigen oncogene under the transcriptional control of regulatory elements from the rat tyrosine hydroxylase gene (Suri et al., 1993). CATH.a cells express several differentiated neuronal characteristics including medium and light chain neurofilament proteins, synaptophysin, tyrosine hydroxylase, and dopamine beta-hydroxylase; they synthesize dopamine and norepinephrine. Conversely, they do not express glial-specific fibrillary acidic protein. To establish definitively that CATH.a cells are of neuronal origin, we characterized the repertoire of voltage-gated inward currents expressed by CATH.a cells. Such inward currents are necessary for neuronal excitability. We report that all CATH.a cells possess a tetrodotoxin-sensitive sodium current (peak amplitude = 590 +/- 319 pA) and 68% possess a high voltage-activated calcium current (peak amplitude = 175 +/- 67 pA). Pharmacological analyses suggest that individual cells express varying levels of L- and N-type calcium current, but no P-type current. In addition, in 55% of the cells with a calcium current, about a half of this current is resistant to selective antagonists for L- and N-type currents, suggesting that another calcium current exists in these CATH.a cells which is not L-, N-, or P-type. The heterogeneous pattern of current detected persisted in several CATH. a subclones, suggesting that factors other than genetic variability influence current expression. The demonstration that CATH.a cells express these currents indicates that they have excitable membrane properties characteristic of neurons. Although many peripheral nervous system (PNS) cell lines exist, very few CNS cell lines with differentiated neuronal properties exist. Since the CATH.a cells can be grown continuously in large amounts, they may be useful for purifying, characterizing, and/or cloning various neuronal-specific molecules and thereby may add to our understanding of CNS catecholaminergic neurons.
Transcripts encoded by 2 different rat genomic clones, rg13 and rg100, appear to be typical brain-specific polyA- RNAs, as defined by previous criteria (rare, polysomal, and postnatally expressed from single-copy genes). However, we have found by using a sensitive nuclease protection assay that low levels of these transcripts (10% and 3%, respectively) are detected in polyA+ RNA. To determine if rg transcripts that fractionate as polyA- could have resulted from nicking of polyA+ RNA, we assessed the integrity of 2 known polyA+ RNAs, those of tyrosine hydroxylase, a 2-kilobase (kb) mRNA, and sodium channel, a 9.5-kb RNA. Using RNA prepared by several different procedures, including LiCl-urea and guanidine thiocyanate followed by CsCl centrifugation, the shorter message fractionated as polyA+ after 2 cycles over oligodeoxythymidine (oligo-dT) cellulose, whereas the majority of the longer sodium channel RNA fractionated as polyA, as assayed by nuclease protection using probes from the 5' end of the 2 genes. However, on Northern blots, the same RNA preparations showed an intact 9.5-kb sodium channel band only in polyA+ RNA, suggesting that the polyA- RNAs were randomly cleaved, resulting in a smear of sizes that could not be detected as a discrete band. These data imply that long messages may be nicked during standard isolation procedures and that this would not be detected by Northern blot analysis.(ABSTRACT TRUNCATED AT 250 WORDS)
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