With the emerging interest in the central effects of OT on affiliative behavior, attention should be directed in understanding the distribution and modes of release of OT within brain regions regulating affiliative behavior. While numerous studies have documented the release of OT within the PVN and SON reviewed in [119
], little attention has been given to the source and mode of release of OT within structures that modulate social behavior, such as the NAcc. Here we briefly describe the brain OT system, including forebrain OT projections, and speculate on the origin of behaviorally relevant OT. We will focus on the prairie vole system where appropriate.
The PVN and SON of the hypothalamus are the main sites of OT production in the brain. In the rat, there are two types of OT neurons present in the PVN, large magnocellular neurons and smaller parvocellular neurons, which differ not only with respect to size, but also with regard to their projections [121
]. The SON contains only magnocellular OT cells [121
]. In prairie voles, OT neurons can also be seen in the medial preotic nucleus, median preoptic nucleus, and preoptic paraventricular nucleus; all which are continuous with the PVN population. Additionally, individual OT labeled neurons can be found in the bed nucleus of the stria terminalis and the lateral hypothalamic area [123
]. Although the soma of OT neurons are mainly restricted to the hypothalamus, OT fibers are spread throughout the entire brain. Sparse, large caliber fibers can be found in the NAcc, amygdala, lateral septum and hippocampus (, also see Supplementary Figure 1
). In the rat, one of the densest central OT projections is to the brainstem and spinal cord [122
Figure 5 Light micrograph of oxytocin-immunoreactive fibers in the prairie vole from a horizontal section. Notice that a few fibers deviate from the neurohypophysial pathway of the paraventricular nucleus of the hypothalamus (PVN) and project toward the nucleus (more ...)
These dense OT fibers in caudal brain areas caught the attention of early researchers. Tracer experiments were done to determine if individual PVN cells project to the brainstem and spinal cord, to the pituitary, or both. It was found that magnocellular cells of the PVN project to the posterior pituitary; while the smaller parvocellular cells project to the hind brain or spinal cord, with 0.2% projecting to both the pituitary and brainstem [124
]. Since these studies, it has been assumed by many in the field that all the central OT fibers originate from parvocellular cells of the PVN, creating a dissociation between the centrally projecting OT system and the neurohypophyseal system. This separation of projection targets is supported by microdialysis studies showing that peripheral and central release are dissociated under certain circumstances [128
Unlike classical neurotransmitters, which are released primarily at the synaptic cleft, neuropeptide neurons can release peptide from its entire surface area [131
] and can diffuse through the extracellular space due to long half-lives. Peptide specificity is achieved through a high binding affinity for the receptors, about 1000× higher than classical neurotransmitters [119
]. Dendritic release of OT has been well characterized and is independent of neuronal firing [for review see [120
released from intracellular stores favors dendritic release, while Ca++
influx from ion channels favors axonal release. Recent studies have begun to elucidate the mechanism by which dendritic release occurs. First, glutamate can trigger dendritic release of OT through the activation of NMDA receptors without producing action potentials [120
]. Alpha melanocyte stimulating hormone (αMSH) evokes dendritic release of OT and inhibits axonal release in the SON. OT neurons in the PVN and SON express the MC4 receptor that binds αMSH and triggers intracellular calcium release to evoke dendritic OT release. At the same time, αMSH inhibits electrical activity and reduces OT release from axons by releasing endocannabinoids to presynaptically inhibit afferent glutamate release [[133
], for review see [120
]]. It has been speculated that dendritically released OT from the SON or PVN diffuses to distant brain regions where it activates OTRs mediating social behavior [119
]. However, little attention has been given to the release of OT from the sparse network of OT fibers coursing through forebrain limbic regions.
As OTR in the NAcc plays a critical role in regulating alloparental behavior and partner preference formation in prairie voles, we have performed a detailed characterization of OT-immunoreactive fibers in the NAcc. The density of OTR binding in the NAcc is highly species specific, with prairie voles having high densities of receptors throughout the striatum, rats having intermediate receptor binding, and mice and meadow voles having little or no OTR binding in the Nacc (). This variability in OTR localization may provide a mechanism by which evolution acts to change social systems from one species to another, as seen between the monogamous and polygamous voles.
Figure 6 Oxytocin receptor binding (A-C) and oxytocin-immunoreactive fiber distribution (D-F) in rats (top), mice (middle) and prairie voles (bottom). Note the remarkable species differences in oxytocin receptor binding in the nucleus accumbens (NAcc), but similarity (more ...)
Since the expression of OTR varies across species, particularly in the NAcc, we were interested in knowing whether the presence of OT fibers in this area also varied concordantly. Surprisingly similar densities of OT-immunoreactive fibers are found in the NAcc of rats, mice, meadow voles, and prairie voles [59
] (). A recent study in the naked mole rat has shown OT fibers are present in the NAcc of this eusocial species as well [136
]. Thus it appears that in contrast to the localization of OTR, the distribution of OT fibers is remarkably conserved across species. Although not shown in a photomicrograph, OT fibers in the NAcc have been mentioned previously in rats using immunohistochemisry [122
] and radioimmunoassay [137
]. However, there have been no studies that have investigated the origin on these NAcc OT-immunoreactive fibers in any species.
The ultrastructure of these accumbal OT fibers has revealed that they are thick unmyelinated processes filled with dense core vesicles. A minority of these fibers form synapses (23%) and terminate mainly on spines (83%) and to a lesser extent on dendritic shafts (17%) [59
]. This is in accordance with OT synapses in other areas of the brain which showed a preference for dendritic over axonal contacts [138
]. Interestingly, the synaptic boutons of the OT-immunoreactive fibers were devoid of dense core vesicles. These synapses make asymmetric contacts and therefore appear to be glutamatergic. However, the majority of OT-immunoreactive elements in the NAcc are packed with OT-immunoreactive vesicles and do not posses synapses [59
]. This suggests that OT may be released from the fiber surface in an en passant
manner. From our studies, it is not possible to determine conclusively whether these processes are axonal or dendritic in nature. However, the presence of synapses associated with some terminals suggests that at least some fibers are axonal projections.
To identify the source of the OT fibers to the forebrain, we injected the retrograde tracer fluorogold (FG) into the NAcc of female prairie voles. We found that OT+ neurons that were also FG+ were only present in the PVN and the SON [59
]. The SON was unexpected since it contains only magnocellular cells that are thought to project exclusively to the posterior pituitary [122
]. However, there is evidence to the contrary. Lesions of the PVN and SON effect different populations of OT fibers, using radioimmunoassay [139
]. Important for our investigation, both types of lesions produced a loss in caudate OT content. A study injecting 3H-leucine into the SON found multiple extrahypothalamic projections, including olfactory bulb, amygdala, cingulum, and locus coeruleus [140
To determine if the OT-immunoreactive fibers are coming from pituitary-projecting cells, we injected FG peripherally; where it can be taken up by regions outside the blood brain barrier, such as the pituitary. In the prairie voles, the majority (~96%) of OT cells in the anterior and medial PVN contained FG. In the posterior PVN there was a dorsal group of smaller OT cells that remained unlabelled, implying that these are the parvocellular cells that project to the hindbrain [59
]. In addition, the PVN cells retrogradely labeled from the NAcc were never seen in this parvocellular population, again suggesting that the OT fibers in the NAcc are coming from magnocellular soma in the PVN.