SELECTIVE DNA SYNTHESIS BY CELLS SPECIFICALLY LOCALIZING IN RESPONSE TO XENOGENEIC ERYTHROCYTES

The present studies have shown that cells capable of specific localization in response to challenge with CRBC or SRBC synthesize DNA very rapidly during the period from 2–5 days (peak 3 days) post primary immunization. This has been done by incubating the antigenically stimulated lymphoid cells with [3H] or [14C]thymidine in vitro for 45 min before adoptive transfer to syngeneic recipients. Specifically localizing cells (SLC) labeled in this way may ultimately account for up to 50% of the 3H or 14C present in a set of specifically challenged lymph nodes 3 days later. The data presented are consistent with the hypothesis that SLC numerically constitute only a very small fraction of the total number of recirculating lymphocytes trapped in antigenically stimulated lymph nodes, and that the demonstration of specific localization therefore depends upon selectively labeling these SLC relative to other recirculating cells. Attempts to selectively label the RNA of SLC with the precursor uridine have to date met with only very limited success

THE IMMUNOLOGICALLY SPECIFIC RETENTION OF RECIRCULATING LONG-LIVED LYMPHOCYTES IN LYMPH NODES STIMULATED BY XENOGENEIC ERYTHROCYTES

The lymph nodes of mice actively or adoptively immunized to sheep RBC and/or chicken RBC selectively retain long-lived lymphocytes after challenge with the appropriate antigen. This retention is demonstrable within 8 hr of the time of stimulation, though it probably begins even before this, and it is essentially complete within the first 24 hr. A similar selective retention is seen in nodes regional to the injection of some nonimmunogenic substances such as turpentine, but not others such as colloidal carbon or syngeneic RBC. In animals adoptively immunized to sheep and chicken RBC simultaneously, there is a preferential accumulation of the labeled long-lived lymphocytes of donors immunized to sheep RBC in lymph nodes challenged with sheep RBC, and a preferential accumulation of lymphocytes (labeled with a different radioisotope) from donors immunized to chicken RBC in lymph nodes challenged with this antigen. This immunologically specific component is demonstrable whether the antigen is given before or after adoptive immunization, suggesting that the only labeled cells capable of specific localization in this system are those cells that normally remain in the recirculating pool. In the present experiments, 31 out of 31 sets of antigenically stimulated lymph nodes have shown radiochemical evidence of immunological specificity in the distribution of donor lymphocytes between them, while corresponding sets of nonstimulated lymph nodes have shown only small random variations in the distribution of donor cells. Two different mechanisms are postulated whereby antigenic stimulation can alter the traffic of recirculating long-lived lymphocytes through stimulated lymph nodes. One affects recirculating cells of a particular immunological specificity, while the other affects recirculating cells without regard to their immunological specificity

CGRPα-Expressing Sensory Neurons Respond to Stimuli that Evoke Sensations of Pain and Itch

Calcitonin gene-related peptide (CGRPα, encoded by Calca) is a classic marker of nociceptive dorsal root ganglia (DRG) neurons. Despite years of research, it is unclear what stimuli these neurons detect in vitro or in vivo. To facilitate functional studies of these neurons, we genetically targeted an axonal tracer (farnesylated enhanced green fluorescent protein; GFP) and a LoxP-stopped cell ablation construct (human diphtheria toxin receptor; DTR) to the Calca locus. In culture, 10–50% (depending on ligand) of all CGRPα-GFP-positive (+) neurons responded to capsaicin, mustard oil, menthol, acidic pH, ATP, and pruritogens (histamine and chloroquine), suggesting a role for peptidergic neurons in detecting noxious stimuli and itch. In contrast, few (2.2±1.3%) CGRPα-GFP+ neurons responded to the TRPM8-selective cooling agent icilin. In adult mice, CGRPα-GFP+ cell bodies were located in the DRG, spinal cord (motor neurons and dorsal horn neurons), brain and thyroid—reproducibly marking all cell types known to express Calca. Half of all CGRPα-GFP+ DRG neurons expressed TRPV1, ∼25% expressed neurofilament-200, <10% contained nonpeptidergic markers (IB4 and Prostatic acid phosphatase) and almost none (<1%) expressed TRPM8. CGRPα-GFP+ neurons innervated the dorsal spinal cord and innervated cutaneous and visceral tissues. This included nerve endings in the epidermis and on guard hairs. Our study provides direct evidence that CGRPα+ DRG neurons respond to agonists that evoke pain and itch and constitute a sensory circuit that is largely distinct from nonpeptidergic circuits and TRPM8+/cool temperature circuits. In future studies, it should be possible to conditionally ablate CGRPα-expressing neurons to evaluate sensory and non-sensory functions for these neurons

Immunologically Specific Retention of Long-Lived Lymphoid Cells in Antigenically Stimulated Lymph Nodes

Abstract The lymphoid cells selectively retained in lymph nodes that have been stimulated by xenogenic erythrocytes include cells of the long-lived subpopulation. This selective retention appears to be real and not an artifact resulting from the reutilization of radioactively labeled DNA or its split products. Lymph nodes draining skin allografts also show a selective retention of long-lived lymphoid cells, although this retention is of a lesser magnitude and occurs at a somewhat slower rate. While most of this selective retention is related to factors other than the immunologic specificity of the long-lived cells involved, we have demonstrated that a small portion of it reflects the accumulation of specific memory cells. This has been done by using an experimental design based on: a) the adoptive immunization of mice to two different antigens (BALB/c mice immunized to DBA/2 and C57BL/Ks histocompatibility antigens), b) the labeling of long-lived lymphocytes of the donor mice with either 3H-thymidine or 14C-thymidine following immunization, and c) the simultaneous counting of both radioisotopes in each set of stimulated lymph nodes.</jats:p