A Corticostriatal Path Targeting Striosomes Controls Decision-Making under Conflict.

A striking neurochemical form of compartmentalization has been found in the striatum of humans and other species, dividing it into striosomes and matrix. The function of this organization has been unclear, but the anatomical connections of striosomes indicate their relation to emotion-related brain regions, including the medial prefrontal cortex. We capitalized on this fact by combining pathway-specific optogenetics and electrophysiology in behaving rats to search for selective functions of striosomes. We demonstrate that a medial prefronto-striosomal circuit is selectively active in and causally necessary for cost-benefit decision-making under approach-avoidance conflict conditions known to evoke anxiety in humans. We show that this circuit has unique dynamic properties likely reflecting striatal interneuron function. These findings demonstrate that cognitive and emotion-related functions are, like sensory-motor processing, subject to encoding within compartmentally organized representations in the forebrain and suggest that striosome-targeting corticostriatal circuits can underlie neural processing of decisions fundamental for survival.

Single-neuron sequencing analysis of L1 retrotransposition and somatic mutation in the human brain.

A major unanswered question in neuroscience is whether there exists genomic variability between individual neurons of the brain, contributing to functional diversity or to an unexplained burden of neurological disease. To address this question, we developed a method to amplify genomes of single neurons from human brains. Because recent reports suggest frequent LINE-1 (L1) retrotransposition in human brains, we performed genome-wide L1 insertion profiling of 300 single neurons from cerebral cortex and caudate nucleus of three normal individuals, recovering >80% of germline insertions from single neurons. While we find somatic L1 insertions, we estimate <0.6 unique somatic insertions per neuron, and most neurons lack detectable somatic insertions, suggesting that L1 is not a major generator of neuronal diversity in cortex and caudate. We then genotyped single cortical cells to characterize the mosaicism of a somatic AKT3 mutation identified in a child with hemimegalencephaly. Single-neuron sequencing allows systematic assessment of genomic diversity in the human brain.

A single dose of escitalopram blunts the neural response in the thalamus and caudate during monetary loss.

Background: Selective serotonin reuptake inhibitors (SSRIs) show acute effects on the neural processes associated with negative affective bias in healthy people and people with depression. However, whether and how SSRIs also affect reward and punishment processing on a similarly rapid time scale remains unclear. Methods: We investigated the effects of an acute and clinically relevant dose (20 mg) of the SSRI escitalopram on brain response during reward and punishment processing in 19 healthy participants. In a doubleblind, placebo-controlled study using functional MRI, participants performed a well-established monetary reward task at 3 time points: at baseline; after receiving placebo or escitalopram; and after receiving placebo or escitalopram following an 8-week washout period. Results: Acute escitalopram administration reduced blood-oxygen-level-dependent (BOLD) response during punishment feedback in the right thalamus (family-wise error corrected [FWE] p = 0.013 at peak level) and the right caudate head (pFWE = 0.011 at peak level) compared to placebo. We did not detect any significant BOLD changes during reward feedback. Limitations: We included only healthy participants, so interpretation of findings are limited to the healthy human brain and require future testing in patient populations. The paradigm we used was based on monetary stimuli, and results may not be generalizable to other forms of reward. Conclusion: Our findings extend theories of rapid SSRI action on the neural processing of rewarding and aversive stimuli and suggest a specific and acute effect of escitalopram in the punishment neurocircuitry.

Propofol rather than Isoflurane Accelerates the Interstitial Fluid Drainage in the Deep Rat Brain.

Objective: Different anesthetics have distinct effects on the interstitial fluid (ISF) drainage in the extracellular space (ECS) of the superficial rat brain, while their effects on ISF drainage in the ECS of the deep rat brain still remain unknown. Herein, we attempt to investigate and compare the effects of propofol and isoflurane on ECS structure and ISF drainage in the caudate-putamen (CPu) and thalamus (Tha) of the deep rat brain. Methods: Adult Sprague-Dawley rats were anesthetized with propofol or isoflurane, respectively. Twenty-four anesthetized rats were randomly divided into the propofol-CPu, isoflurane-CPu, propofol-Tha, and isoflurane-Tha groups. Tracer-based magnetic resonance imaging (MRI) and fluorescent-labeled tracer assay were utilized to quantify ISF drainage in the deep brain. Results: The half-life of ISF in the propofol-CPu and propofol-Tha groups was shorter than that in the isoflurane-CPu and isoflurane-Tha groups, respectively. The ECS volume fraction in the propofol-CPu and propofol-Tha groups was much higher than that in the isoflurane-CPu and isoflurane-Tha groups, respectively. However, the ECS tortuosity in the propofol-CPu and propofol-Tha groups was much smaller than that in isoflurane-CPu and isoflurane-Tha groups, respectively. Conclusions: Our results demonstrate that propofol rather than isoflurane accelerates the ISF drainage in the deep rat brain, which provides novel insights into the selective control of ISF drainage and guides selection of anesthetic agents in different clinical settings, and unravels the mechanism of how general anesthetics function.

Convergence of prefrontal and parietal anatomical projections in a connectional hub in the striatum.

Visual attentional bias forms for rewarding and punishing stimuli in the environment. While this attentional bias is adaptive in healthy situations, it is maladaptive in disorders such as drug addiction or PTSD. In both these disorders, the ability to exert control over this attentional bias is associated with drug abstinence rates or reduced PTSD symptoms, indicating the interaction of visual attention, cognitive control, and stimulus association. The inferior parietal lobule (IPL) is central to attention, while the prefrontal cortex (PFC) is critical for reward, cognitive control, and attention. Importantly, regions of the IPL and PFC commonly project to the rostral dorsal caudate (rdCaud) of the striatum. We propose an anatomical network architecture in which IPL projections converge with PFC projections in a connectional hub in the rdCaud, providing an anatomical substrate for the interaction of these projections and their competitive influence on striatal processing. To investigate this, we mapped the dense projections from the caudal IPL and prefrontal (dlPFC, vlPFC, OFC, dACC, and dmPFC) regions that project to the medial rdCaud with anatomical tract-tracing tracer injections in monkeys. These inputs converge in a precise site in the medial rdCaud, rostral to the anterior commissure. Small retrograde tracer injections confirmed these inputs to the medial rdCaud and showed that a proximal ventral striatal location has a very different pattern of cortical inputs. We next used human resting-state functional connectivity MRI (fcMRI) to examine whether a striatal hub exists in the human medial rdCaud. Seed regions in the human medial rdCaud revealed cortical correlation maps similar to the monkey retrograde injection results. A subsequent analysis of these correlated cortical regions showed that their peak correlation within the striatum is in the medial rdCaud, indicating that this is a connectional hub. In contrast, this peak striatal correlation was not found in the ventral striatal location, suggesting that this site is not a connectional hub of cortical regions. Taken together, this work uses the precision of monkey anatomy to identify a connectional hub of IPL and PFC projections in the medial rdCaud. It also translates this anatomical precision to humans, demonstrating that, guided by anatomy, connectional hubs can be identified in humans with fcMRI. These connectional hubs provide more specific treatment targets for drug addiction, PTSD, and other neurological and psychiatric disorders involving the striatum.

Projections from caudal ventrolateral prefrontal areas to brainstem preoculomotor structures and to Basal Ganglia and cerebellar oculomotor loops in the macaque.

The caudal part of the macaque ventrolateral prefrontal (VLPF) cortex hosts several distinct areas or fields--45B, 45A, 8r, caudal 46vc, and caudal 12r--connected to the frontal eye field (area 8/FEF). To assess whether these areas/fields also display subcortical projections possibly mediating a role in controlling oculomotor behavior, we examined their descending projections, based on anterograde tracer injections in each area/field, and compared them with those of area 8/FEF. All the studied areas/fields displayed projections to brainstem preoculomotor structures, precerebellar centers, and striatal sectors that are also targets of projections originating from area 8/FEF. Specifically, these projections involved: (1) the intermediate and superficial layers of the superior colliculus; (2) the mesencephalic and pontine reticular formation; (3) the dorsomedial and lateral pontine nuclei and the reticularis tegmenti pontis; and (4) the body of the caudate nucleus. Furthermore, area 45B projected also to the regions around the trochlear nucleus and to the raphe interpositus. The present data provide evidence for a role of the caudal VLPF areas/fields in controlling oculomotor behavior not only through their connections to area 8/FEF, but also in parallel through a direct access to preoculomotor brainstem structures and to the cerebellar and basal ganglia oculomotor loops.

Representation of outcome risk and action in the anterior caudate nucleus.

The anterior caudate nucleus is essential for goal-directed behavior because it links outcome information to actions. It is well known that caudate neurons provide a variety of reward-related and action signals. However, it is still unclear how the two signals are integrated. We investigated whether and how outcome risk modulates spatial representation. We recorded neural activity in the anterior caudate nucleus while monkeys made saccades to multiple spatial targets, each associated with either fixed (safe) or variable (risky) amount of reward. We report that individual neurons combined the outcome reward signal with spatial information about the direction of saccades. These signals could be reliably read out from the populations of neurons. Moreover, the prospect of a risky outcome improved the quality of spatial information. These results provide direct evidence that global spatial representation in the caudate is modulated by outcome, which can be important for flexible control of behavior, particularly during learning and habit formation, when outcomes vary.

Ovarian steroids alter dopamine receptor populations in the medial preoptic area of female rats: implications for sexual motivation, desire, and behaviour.

Dopamine (DA) transmission in the medial preoptic area (mPOA) plays a critical role in the control of appetitive sexual behaviour in the female rat. We have shown previously that a DA D1 receptor (D1R)-mediated excitatory state appears to occur in females primed with estradiol benzoate (EB) and progesterone (P), whereas a DA D2 receptor (D2R)-mediated inhibitory state appears to occur in females primed only with EB. The present experiment employed three techniques to better understand what changes occur to DA receptors (DARs) in the mPOA under different hormonal profiles. Ovariectomized females were randomly assigned to one of three steroid treatment groups: EB + P (10 and 500 µg, respectively), EB + Oil, or the control (Oil + Oil), with hormone injections administered at 48 and 4 h prior to euthanizing. First, the number of neurons in the mPOA that contained D1R or D2R was assessed using immunohistochemistry. Second, the mPOA and two control areas (the prelimbic cortex and caudate putamen) were analysed for DAR protein levels using western blot, and DAR functional binding levels using autoradiography. Ovarian steroid hormones affected the two DAR subtypes in opposite ways in the mPOA. All three techniques supported previous behavioural findings that females primed with EB have a lower D1R : D2R ratio, and thus a D2R-mediated system, and females primed with EB + P have a higher D1R : D2R ratio, and thus a D1R-mediated system. This provides strong evidence for a DA-driven pathway of female sexual motivation, desire, and behaviour that is modified by different hormone priming regimens.

Interactions between glutamate, dopamine, and the neuronal signature of response inhibition in the human striatum.

Response inhibition is a basic mechanism in cognitive control and dysfunctional in major psychiatric disorders. The neuronal mechanisms are in part driven by dopamine in the striatum. Animal data suggest a regulatory role of glutamate on the level of the striatum. We used a trimodal imaging procedure of the human striatum including F18-DOPA positron emission tomography, proton magnetic resonance spectroscopy, and functional magnetic resonance imaging of a stop signal task. We investigated dopamine synthesis capacity and glutamate concentration in vivo and their relation to functional properties of response inhibition. A mediation analysis revealed a significant positive association between dopamine synthesis capacity and inhibition-related neural activity in the caudate nucleus. This relationship was significantly mediated by striatal glutamate concentration. Furthermore, stop signal reaction time was inversely related to striatal activity during inhibition. The data show, for the first time in humans, an interaction between dopamine, glutamate, and the neural signature of response inhibition in the striatum. This finding stresses the importance of the dopamine-glutamate interaction for behavior and may facilitate the understanding of psychiatric disorders characterized by impaired response inhibition.

A model system for in vivo gene transfer into the central nervous system using an adenoviral vector.

Previous methods of in vivo gene transfer to differentiated neurons of the adult mammalian brain have been inefficient and associated with technical problems. We have therefore developed a model system of direct gene transfer using a replication-defective adenoviral vector containing a beta-galactosidase gene to transduce brain neurons. Following injection of purified high titre recombinant adenovirus into the caudate putamen of seven week old mice, lacZ activity was evident in neural components of the central nervous system (CNS) for at least 8 weeks post infection. The efficiency of adenoviral gene transfer was very high compared to other techniques, suggesting an attractive and efficient alternative for neuronal gene transfer in vivo.

Encoding of both positive and negative reward prediction errors by neurons of the primate lateral prefrontal cortex and caudate nucleus.

Learning can be motivated by unanticipated success or unexpected failure. The former encourages us to repeat an action or activity, whereas the latter leads us to find an alternative strategy. Understanding the neural representation of these unexpected events is therefore critical to elucidate learning-related circuits. We examined the activity of neurons in the lateral prefrontal cortex (PFC) and caudate nucleus of monkeys as they performed a trial-and-error learning task. Unexpected outcomes were widely represented in both structures, and neurons driven by unexpectedly negative outcomes were as frequent as those activated by unexpectedly positive outcomes. Moreover, both positive and negative reward prediction errors (RPEs) were represented primarily by increases in firing rate, unlike the manner in which dopamine neurons have been observed to reflect these values. Interestingly, positive RPEs tended to appear with shorter latency than negative RPEs, perhaps reflecting the mechanism of their generation. Last, in the PFC but not the caudate, trial-by-trial variations in outcome-related activity were linked to the animals' subsequent behavioral decisions. More broadly, the robustness of RPE signaling by these neurons suggests that actor-critic models of reinforcement learning in which the PFC and particularly the caudate are considered primarily to be "actors" rather than "critics," should be reconsidered to include a prominent evaluative role for these structures.

Saccade reaction times are influenced by caudate microstimulation following and prior to visual stimulus appearance.

Several cognitive models suggest that saccade RTs are controlled flexibly not only by mechanisms that accumulate sensory evidence after the appearance of a sensory stimulus (poststimulus mechanisms) but also by mechanisms that preset the saccade control system before the sensory event (prestimulus mechanisms). Consistent with model predictions, neurons in structures tightly related to saccade initiation, such as the superior colliculus and FEF, have poststimulus and prestimulus activities correlated with RTs. It has been hypothesized that the BG influence the saccade initiation process by controlling both poststimulus and prestimulus activities of superior colliculus and FEF neurons. To examine this hypothesis directly, we delivered electrical microstimulation to the caudate nucleus, the input stage of the oculomotor BG, while monkeys performed a prosaccade (look toward a visual stimulus) and antisaccade (look away from the stimulus) paradigm. Microstimulation applied after stimulus appearance (poststimulus microstimulation) prolonged RTs regardless of saccade directions (contra/ipsi) or task instructions (pro/anti). In contrast, microstimulation applied before stimulus appearance (prestimulus microstimulation) shortened RTs, although the effects were limited to several task conditions. The analysis of RT distributions using the linear approach to threshold with ergodic rate model revealed that poststimulus microstimulation prolonged RTs by reducing the rate of rise to the threshold for saccade initiation, whereas fitting results for prestimulus microstimulation were inconsistent across different task conditions. We conclude that both poststimulus and prestimulus activities of caudate neurons are sufficient to control saccade RTs.

HIV-1 gp120 upregulates matrix metalloproteinases and their inhibitors in a rat model of HIV encephalopathy.

Matrix metalloproteinases (MMPs) are implicated in diverse processes, such as neuroinflammation, leakiness of the blood-brain barrier (BBB) and direct cellular damage in neurodegenerative and other CNS diseases. Tissue destruction by MMPs is regulated by their endogenous tissue inhibitors (TIMPs). TIMPs prevent excessive MMP-related degradation of extracellular matrix components. In a rat model of human immunodeficiency virus (HIV)-related encephalopathy, we described MMP-2 and MMP-9 upregulation by HIV-1 envelope gp120, probably via gp120-induced reactive oxygen species. Antioxidant gene delivery blunted gp120-induced MMP production. We also studied the effect of gp120 on TIMP-1 and TIMP-2 production. TIMP-1 and TIMP-2 levels increased 6 h after gp120 injection into rat caudate-putamen (CP). TIMP-1 and TIMP-2 colocalized mainly with neurons (92 and 95%, respectively). By 24 h, expression of these protease inhibitors diverged, as TIMP-1 levels remained high but TIMP-2 subsided. Gene delivery of the antioxidant enzymes Cu/Zn superoxide dismutase or glutathione peroxidase into the CP before injecting gp120 there reduced levels of gp120-induced TIMP-1 and TIMP-2, recapitulating the effect of antioxidant enzymes on gp120-induced MMP-2 and MMP-9. A significant correlation was observed between MMP/TIMP upregulation and BBB leakiness. Thus, HIV-1 gp120 upregulated TIMP-1 and TIMP-2 in the CP. Prior antioxidant enzyme treatment mitigated production of these TIMPs, probably by reducing MMP expression.

Neuroprotective effects of phytocannabinoid-based medicines in experimental models of Huntington's disease.

We studied whether combinations of botanical extracts enriched in either Δ(9)-tetrahydrocannabinol (Δ(9)-THC) or cannabidiol (CBD), which are the main constituents of the cannabis-based medicine Sativex, provide neuroprotection in rat models of Huntington's disease (HD). We used rats intoxicated with 3-nitropropionate (3NP) that were given combinations of Δ(9)-THC- and CBD-enriched botanical extracts. The issue was also studied in malonate-lesioned rats. The administration of Δ(9)-THC- and CBD-enriched botanical extracts combined in a ratio of 1:1 as in Sativex attenuated 3NP-induced GABA deficiency, loss of Nissl-stained neurons, down-regulation of CB(1) receptor and IGF-1 expression, and up-regulation of calpain expression, whereas it completely reversed the reduction in superoxide dismutase-1 expression. Similar responses were generally found with other combinations of Δ(9)-THC- and CBD-enriched botanical extracts, suggesting that these effects are probably related to the antioxidant and CB(1) and CB(2) receptor-independent properties of both phytocannabinoids. In fact, selective antagonists for both receptor types, i.e., SR141716 and AM630, respectively, were unable to prevent the positive effects on calpain expression caused in 3NP-intoxicated rats by the 1:1 combination of Δ(9)-THC and CBD. Finally, this combination also reversed the up-regulation of proinflammatory markers such as inducible nitric oxide synthase observed in malonate-lesioned rats. In conclusion, this study provides preclinical evidence in support of a beneficial effect of the cannabis-based medicine Sativex as a neuroprotective agent capable of delaying disease progression in HD, a disorder that is currently poorly managed in the clinic, prompting an urgent need for clinical trials with agents showing positive results in preclinical studies.

Morphometric analysis of neuronal and glial cell pathology in the caudate nucleus in late-life depression.

OBJECTIVE: To assess glial and neuronal density and neuronal volume in two areas of the caudate nucleus in late-life major depression. DESIGN: A postmortem study using the disector and nucleator methods to estimate neuronal density and volume and glial density of cells from human brain tissue from the anterior portion (dorsolateral and ventromedial aspects) of the caudate nucleus. SETTING: Brain tissues were obtained from the Newcastle Brain Tissue Resource at Newcastle University, UK. PARTICIPANTS: The study group consisted of 13 subjects with late-life major depression and nine comparison subjects of similar age. RESULTS: Evidence of moderate reductions in neuronal density was found in the depressed group in both the dorsolateral and ventromedial areas of the caudate nucleus. There were no significant changes in glial density or neuronal volume in either area nor was there any evidence of differences in depression in early and late-onset subgroups. CONCLUSIONS: Neuroimaging abnormalities in frontal and subcortical areas including ischemic hyperintensities and a reduction in volume and metabolism in the caudate nucleus have been reported in late-life depression, and previous morphometric studies have reported neuronal changes in prefrontal cortical areas. The findings in this study extend these morphometric investigations in late-life depression to the caudate nucleus, suggesting that neuronal abnormalities are present in this subcortical nucleus as well as in these related prefrontal areas.

Selective enhancement of associative learning by microstimulation of the anterior caudate.

Primates have the remarkable ability to rapidly adjust or modify associations between visual cues and specific motor responses. Whereas little is known as to how such adjustments in behavioral policy are implemented, recent learning models suggest that the anterior striatum is optimally positioned to have a role in this process. We recorded from single units and delivered microstimulation in the striatum of rhesus monkeys performing an associative learning task. Caudate activity during reinforcement was closely correlated with the rate of learning and peaked during the steepest portion of the learning curve when new associations were being acquired. Moreover, delivering microstimulation in the caudate during the reinforcement period significantly increased the rate of learning without altering the monkeys' ultimate performance. These findings suggest that the caudate is responsible for implementing selective adjustments to the 'associative weights' between sensory cues and motor responses during learning, thus enhancing the likelihood of selecting profitable actions.

[Imaging and quantitative measurement of brain extracellular space using MRI Gd-DTPA tracer method].

OBJECTIVE: To observe the diffusion of Gd-DTPA in brain extracellular space (ECS) by magnetic resonance imaging(MRI) and investigate the feasibility of ECS measurement by using MRI tracer method in vivo. METHODS: 2 microL Gd-DTPA was introduced into ECS by caudate nucleus according to stereotaxic atlas in 8 Sprague Dawley(SD) rats (male, 280-320 g). The MRI scans were performed at 1 h, 3 h, 6 h, 9 h and 12 h respectively after administration. MRI appearances of Gd-DTPA diffusion and distribution was observed and compared. The MRI signal enhancement was measured at each time point. The neuroethology assessment was performed after MRI scanning at 12 h. RESULTS: The injection was accurate at the center of the caudate nucleus in 6 rats, while, at the capsula externa in other 2 rats. Gd-DTPA diffused isotropically after it was introduced into caudate nucleus, which spread into lateral cortex at 3 h. The MRI signal enhancement distributed mainly in the middle cerebral artery territory. A significant difference was found between the signal enhancement ratio at 1 h and that at 3 h in the original point of caudate nucleus (t=95.63, P<0.01), and the signal enhancement attenuated following the exponential power function y=1.7886x(-0.1776) (R2=0.94). In 2 rats with the injection point at capsula externa, Gd-DTPA diffused anisotropically along the fiber track of white matter during 1 h to 3 h, and spread into the lateral cortex at 6 h. CONCLUSION: The diffusion and clearance of Gd-DTPA in brain ECS could be monitored and measured quantitatively in vivo by MRI tracer method.

Frontal eye field and caudate neurons make different contributions to reward-biased perceptual decisions.

Many decisions require trade-offs between sensory evidence and internal preferences. Potential neural substrates include the frontal eye field (FEF) and caudate nucleus, but their distinct roles are not understood. Previously we showed that monkeys' decisions on a direction-discrimination task with asymmetric rewards reflected a biased accumulate-to-bound decision process (Fan et al., 2018) that was affected by caudate microstimulation (Doi et al., 2020). Here we compared single-neuron activity in FEF and caudate to each other and to accumulate-to-bound model predictions derived from behavior. Task-dependent neural modulations were similar in both regions. However, choice-selective neurons in FEF, but not caudate, encoded behaviorally derived biases in the accumulation process. Baseline activity in both regions was sensitive to reward context, but this sensitivity was not reliably associated with behavioral biases. These results imply distinct contributions of FEF and caudate neurons to reward-biased decision-making and put experimental constraints on the neural implementation of accumulation-to-bound-like computations.

Reduced density of oligodendrocytes and oligodendrocyte clusters in the caudate nucleus in major psychiatric illnesses.

Functional dysconnectivity in schizophrenia and affective disorders may be associated with myelin and oligodendrocyte abnormalities. Altered network integration involving the caudate nucleus (CN) and metabolic abnormalities in fronto-striatal-thalamic white matter tracts have been reported in schizophrenia and impaired patterns of cortico-caudate functional connectivity have been found in both bipolar disorder (BPD) and schizophrenia compared to healthy controls. Postmortem studies have found ultrastructural dystrophy and degeneration of oligodendrocytes and dysmyelination in the CN in schizophrenia and BPD. We aimed to test the hypothesis that oligodendrocyte density may be reduced in the CN in major psychiatric disorders and may thereby form the cellular basis for the functional dysconnectivity observed in these disorders. Optical disector was used to estimate the numerical density (Nv) of oligodendrocytes and oligodendrocyte clusters (OLC) in the CN of cases with schizophrenia, BPD and major depressive disorder (MDD) and in normal controls (15 cases per group). A significant reduction in the Nv of oligodendrocytes was found in schizophrenia and BPD as compared to the control group (p < 0.05), and the Nv of OLC was significantly lowered in schizophrenia and BPD compared to controls (p < 0.05). There were no significant differences between MDD and control groups. The Nv of OLC was significantly decreased in the left hemisphere in schizophrenia as compared to the left hemisphere of the control group (-52%, p < 0.01). The data indicates that a decreased density of oligodendrocytes and OLC could contribute to the altered functional connectivity of the CN in subjects with severe mental illnesses.

Immunocytochemical localization studies of myelin basic protein.

The location of myelin encephalitogenic or basic protein (BP) in peripheral nervous system (PNS) and central nervous system (CNS) was investigated by immunofluorescence and horseradish peroxidase (HRP) immunocytochemistry. BP or cross-reacting material could be clearly localized to myelin by immunofluorescence and light microscope HRP immunocytochemistry. Fine structural studies proved to be much more difficult, especially in the CNS, due to problems in tissue fixation and penetration of reagents. Sequential fixation in aldehyde followed by ethanol or methanol provided the best conditions for ultrastructural indirect immunocytochemical studies. In PNS tissue, anti-BP was localized exclusively to the intraperiod line of myelin. Because of limitations in technique, the localization of BP in CNS myelin could not be unequivocally determined. In both PNS and CNS tissue, no anti-BP binding to nonmyelin cellular or membranous elements was detected.