Christine J. Harling-Berg

Drainage of Brain Extracellular Fluid into Blood and Deep Cervical Lymph and its Immunological Significance

Cerebral extracellular fluids drain from brain to blood across the arachnoid villi and to lymph along certain cranial nerves (primarily olfactory) and spinal nerve root ganglia. Quantification of the connection to lymph in rabbit, cat and sheep, using radiolabeled albumin as a marker of flow, indicates that a minimum of 14 to 47% of protein injected into different regions of brain or cerebrospinal fluid passes through lymph. The magnitude of the outflow to lymph is at variance with the general assumption that the absence of conventional lymphatics from the brain interrupts the afferent arm of the immune response to brain antigens. The immune response to antigens (albumin or myelin basic protein) introduced into the central nervous system (CNS) has been analysed using a rat model with normal brain barrier permeability. The micro‐injection of antigen into brain or cerebrospinal fluid elicits a humoral immune response, with antibody production in cervical lymph nodes and spleen, and also affects cell‐mediated immunity. Furthermore, antigen may be more immunogenic when administered into the CNS than into conventional extracerebral sites. Clearly, the afferent arm of the immune response to antigens, within the CNS, is intact. Modern studies suggest that the efferent arm is also intact with passage of activated lymphocytes into the brain. Results support a new view of CNS immunology which incorporates continuous and highly regulated communication between the brain and the immune system in both health and disease.

Role of cervical lymph nodes in the systemic humoral immune response to human serum albumin microinfused into rat cerebrospinal fluid.

The humoral immune response to human serum albumin (HSA) microinfused into cerebrospinal fluid (CSF) has been measured in serum, cervical lymph nodes, and spleen of Sprague-Dawley rats. Conditions were designed to promote normal brain barrier function. Serum titers of anti-HSA antibodies, primarily IgG, increased over 10 days and then persisted for at least 10 weeks. A significant role for cervical lymphatics in the systemic response to CSF-administered HSA is suggested, based on results showing that (1) cervical lymph obstruction reduces serum titers of anti-HSA antibodies, and (2) total antibody production by combined superficial and deep cervical nodes, sampled 14 days post-immunization, exceeds that by the spleen.

Anti-striatal antibodies in Tourette syndrome cause neuronal dysfunction

Serologic studies of children with Tourette syndrome (TS) have detected anti-neuronal antibodies but their role in TS has not been explored. Stereotypies and episodic utterances, analogous to involuntary movements seen in TS, were induced in rats by intrastriatal microinfusion of TS sera or gamma immunoglobulins (IgG) under noninflammatory conditions, as found in TS. Immunohistochemical analysis confirmed the presence of IgG selectively bound to striatal neurons. These data support the hypothesis that binding of an anti-neuronal antibody from some children with TS induced striatal dysfunction and suggest a possible cause for the basal ganglia alterations observed in children with TS.

Role of the cervical lymphatics in the Th2-type hierarchy of CNS immune regulation.

CNS immune regulation is intimately dependent on the dynamics of cerebral extracellular fluid circulation. Animal models indicate that following the introduction of antigen into the CNS, normal circulation of interstitial and cerebrospinal fluids provides the opportunity for (a) delivery of CNS-derived antigen to lymphoid organs, as well as, (b) retention of immunologically significant amounts of antigen within the CNS. Thus, even in the absence of disease, CNS-derived antigen can induce antigen-specific activation of naive lymphocytes in lymphoid organs and specific reactivation of lymphoblasts that have migrated into the CNS. The initial peripheral immune response to CNS-derived antigen is induced in cervical lymph nodes and is characterized by a strong antibody response, no delayed-type hypersensitivity, and only priming for cytotoxic T-cell responses. This Th-2 type hierarchy of immune regulation is reinforced within the antigen-stimulated CNS where specific B lymphoblasts are permitted to develop their effector function but cell-mediated immunity is inhibited. Developing a paradigm for CNS immune regulation is important in understanding how CNS disorders in humans are induced, perpetuated, and may be manipulated.

Physiology and immunology of lymphatic drainage of interstitial and cerebrospinal fluid from the brain

Cerebrospinal fluid (CSF) and brain interstitial fluid, extracellular fluids produced in the central nervous system (CNS). efflux by several routes (Figure 1) [reviewed in 4, 51. A major outflow pathway for extracellular fluid is passage through arachnoid villi into blood of the dural sinus. Another efflux pathway is drainage into lymph along routes adjacent to cranial and spinal nerves and leads to regional lymph nodes [15]. Following injection of radiolabelled protein into the CNS, a significant fraction (14-47%) of this tracer leaving the CNS appears in cervical lymph (Table 1). Furthermore, protein injected into the CNS reaches the cervical lymph in a relatively higher concentration compared with protein injected into the blood. Drainage to cervical lymph is dependent upon several factors, including site of injection into brain, animal species, and assay time period (Table 1). Drainage along the olfactory route, through the cribriform plate, and into cervical lymph is a significant pathway in animals [l]. Both blood and lymph efflux pathways are accessible to cells as well as to macromolecules. Thus, passage of these constituents to the spleen and draining lymph nodes raises interesting issues concerning their immunogenicity. Classical observations that allografts survive for more extensive time periods when implanted into brain (compared to peripheral sites) have led to the concept of immune privilege of the brain [2]. In our laboratory, we have developed a model in rats to analyse the immunological significance of antigen outflow from the CNS [8]. An important feature of this model is that a

Role of the cervical lymphatics in the Th2-type hierarchy of CNS immune regulation1

CNS immune regulation is intimately dependent on the dynamics of cerebral extracellular fluid circulation. Animal models indicate that following the introduction of antigen into the CNS, normal circulation of interstitial and cerebrospinal fluids provides the opportunity for (a) delivery of CNS-derived antigen to lymphoid organs, as well as, (b) retention of immunologically significant amounts of antigen within the CNS. Thus, even in the absence of disease, CNS-derived antigen can induce antigen-specific activation of naive lymphocytes in lymphoid organs and specific reactivation of lymphoblasts that have migrated into the CNS. The initial peripheral immune response to CNS-derived antigen is induced in cervical lymph nodes and is characterized by a strong antibody response, no delayed-type hypersensitivity, and only priming for cytotoxic T-cell responses. This Th-2 type hierarchy of immune regulation is reinforced within the antigen-stimulated CNS where specific B lymphoblasts are permitted to develop their effector function but cell-mediated immunity is inhibited. Developing a paradigm for CNS immune regulation is important in understanding how CNS disorders in humans are induced, perpetuated, and may be manipulated.

Afferent and efferent arms of the humoral immune response to CSF-administered albumins in a rat model with normal blood-brain barrier permeability

Cerebrospinal fluid (CSF) and serum antibody responses to albumin administered into CSF or muscle have been compared with respect to titer, isotype profile and complement-fixing activity in a rat model with normal brain barrier function. CSF/serum titer ratios and the ratio of IgG subclasses, IgG1/IgG2, were both elevated following CSF immunization. In contrast, there was no difference in complement-fixing activity between antibodies elicited by the two routes of immunization. It is suggested that intrathecal antibody synthesis accounts for the elevated CSF antibody titers in CSF-immunized rats, providing the first example of central nervous system antibody synthesis in an animal with normal brain barrier permeability.

Myelin basic protein infused into cerebrospinal fluid suppresses experimental autoimmune encephalomyelitis

We have evaluated the antibody and the effector T-cell responses to a single cerebrospinal fluid (CSF) infusion of myelin basic protein (MBP) in Lewis rats by measuring serum anti-MBP antibodies and clinical signs of experimental autoimmune encephalomyelitis (EAE), respectively. Some rats developed anti-MBP antibodies, but none manifested EAE in response to the primary infusion. Antibody responses to an EAE challenge 3 weeks after CSF infusion were normal, but clinical symptoms of EAE were markedly suppressed. Brain trauma at the time of MBP pretreatment enhanced this suppression. The CSF route of MBP administration is more effective in inducing suppression of EAE than peripheral routes.

NORMAL CEREBROSPINAL FLUID SUPPRESSES THE IN VITRO DEVELOPMENT OF CYTOTOXIC T CELLS : ROLE OF THE BRAIN MICROENVIRONMENT IN CNS IMMUNE REGULATION

The regulatory role of cerebrospinal fluid (CSF) in brain physiology is well established, while our understanding of its role in brain immunity is undefined. We demonstrate that normal rat CSF suppresses the in vitro development of mastocytoma-specific CTL activity in restimulated splenocytes from Balb/c mice, a strain unable to reject this tumor from the brain. Suppression is dependent on TGF-beta, revealed by reversal of suppression with specific neutralizing antibody. In contrast, mice which can reject this tumor from the brain, such as Balb/c mice with immunological memory to the tumor or CD-1 mice with major histo-incompatibility with the tumor, have populations of precursor CTL which are resistant to CSF-induced suppression, in the in vitro restimulation protocol. We propose that the susceptibility to CSF-induced suppression of peripherally generated immune cells that traffic to the brain plays an important role in determining whether growing tumor cells survive in the brain.

Hierarchy of immune responses to antigen in the normal brain.

For approximately 100 years the brain has been classified as an “immunologically privileged organ” based on the observations that tissue transplants into cerebral cortex survive longer than tissue transplants into conventional peripheral sites (Shirai 1921; Murphy and Sturm 1923; Medawar 1948; Scheinberg et al. 1966). Various conclusions and presumptions were attributed to these studies, including two predominant beliefs that: (1) there is no lymphatic drainage to alert the immune system to the presence of antigen in the brain (afferent arm of immunity), and (2) the tight endothelial junctions of the cerebral vasculature (blood-brain barrier; BBB) prevent blood lymphocytes from facilitating antigen elimination in the brain (efferent arm of immunity; for reviews see Barker and Billingham 1977; Brent 1990; Waksman 1998).