Sticking out of the crowd: the molecular identity and development of cholecystokinin-containing basket cells

Abstract

Certain essential cognitive processes require the precise temporal interplay between glutamatergic (excitatory) pyramidal cells and γ-aminobutyric acid (GABA)-releasing inhibitory interneurons in the hippocampus. Basket cells, the main class of interneurons, target pyramidal cell somata and proximal dendrites and thus are poised to modify network oscillations. Though only present in limited numbers, the impaired development of basket cells can result in changes in the hippocampal circuitry leading to neurological disorders, such as schizophrenia. The diversity of the spatial origins, neurochemical make-up, cytoarchitecture and network contributions amongst basket cells is a provocative example of interneuron heterogeneity in the hippocampus. This review discusses recent data concerned with the developmental trajectories of one subclass, the cholecystokinin-containing basket cell, and emphasizes the significance of the short-range intercellular guidance cues that have recently emerged to impact the formation and function of their inhibitory synapses.

Tibor Harkany (left) is jointly appointed as professor of Developmental Neurobiology at the Karolinska Institute (Stockholm, Sweden) and SULSA professor of Cell Biology at the University of Aberdeen (UK). After receiving his PhD from the Semmelweis Medical School in Budapest, Hungary, he worked as a post-doctoral fellow with Paul Luiten and Patrik Ernfors in the Netherlands and Sweden, respectively. Erik Keimpema (right) is a recent PhD graduate from Tibor Harkany's lab in Aberdeen who currently works as a post-doctoral fellow at the Karolinska Institute. Harkany and his colleagues investigate the molecular biology of endocannabinoid signalling in the developing brain, particularly in interneurons, with emphasis on the molecular imprint of maternal cannabis abuse in the offspring.

Functional significance of basket cells in the hippocampal circuitry

Some cognitive processes, including attention and memory formation, require the precise temporal interplay between glutamatergic (excitatory) pyramidal cells and γ-aminobutyric acid (GABA)-releasing inhibitory interneurons in the hippocampus (McBain et al. 1999; Freund, 2003; Buzsaki et al. 2004; Morellini et al. 2010; Murray et al. 2011). Although vastly outnumbered by pyramidal cells, the precisely timed activity of interneurons and their innate property to entrain large principal cell ensembles are essential to set the output of the hippocampal circuitry (Freund, 2003; Freund & Katona, 2007).

Interneurons are historically considered as the ‘diverse’ cell type in the hippocampal circuitry, which have evolved to define specific network modalities through selective positioning, spike timing and input–output relationships. The presently accepted consensus to classify interneurons is based on the ‘Petilla’ terminology, which classifies GABAergic interneurons by collating their molecular, anatomical and physiological characteristics (Ascoli et al. 2008). This approach requires multiparametric analysis of the interneurons’ birth places, migratory routes, neurochemical tags, molecular make-up (Table 1), cytoarchitectural features, intrinsic membrane properties and network relationships (Pleasure et al. 2000; Anderson et al. 2002; Ascoli et al. 2008; Tricoire et al. 2011).

Table 1.

Molecular parameters of CCK- and PV-containing cells

CCK+ small basket cell		PV+ nest/large basket cell
	References		References
Receptors		Receptors
5-HT3A	Morales & Bloom, 1997	M2 muscarinic ACh	Hajos et al. 1998
Nicotinic ACh	Porter et al. 1999	μ-Opioid	Drake & Milner, 2002
CB1R	Katona et al. 1999
Oestrogen receptor α	Hart et al. 2007
GABAB	Sloviter et al. 1999
Peptides		Peptides
VIP	Kawaguchi & Kubota, 1997	NPY (subset)	Wang et al. 2002
CRF	Kubota et al. 2011
Endocannabinoids		Endocannabinoids
MGL	Gulyas et al. 2004	MGL	Gulyas et al. 2004
Vesicular transporters		K+ channels
VGLUT3	Somogyi et al. 2004	Kv3.1b	Sekirnjak et al. 1997
		Kv3.2	Hernandez-Pineda et al. 1999
		Kv3.3	Chang et al. 2007

Using these criteria, at least 22 subtypes of GABAergic interneurons are presently discerned in the hippocampus alone. The term ‘basket cell’ alludes to the axonal field of these cells being restricted to the soma and proximal dendrites of principal cells, with their synapses enwrapping excitatory perikarya in the pyramidal layer (Gulyas et al. 1999; Papp et al. 2001). This synapse distribution confers particular power to tune pyramidal cell excitation and network oscillations (Freund, 2003). Basket cells can be divided into neurochemically and functionally distinct classes, the majority expressing parvalbumin (PV) (Nomura et al. 1997), a cytosolic fast Ca²⁺ buffer implicated in the generation and maintenance of high-frequency action potential trains known as ‘fast-spiking’ (>100 Hz). A second class expresses cholecystokinin (CCK), a peptide hormone, and exhibits irregular discharge patterns unusually exceeding 50 Hz (Freund & Buzsaki, 1996; Kawaguchi & Kondo, 2002; Wang et al. 2002). A division of function between PV and CCK basket cells has been proposed with PV⁺ interneurons controlling the rhythm of network oscillations (‘clockwork precision’), while CCK⁺ cells sculpt the efficacy of information flow and encoding by fine-tuning network chronosynchrony (Freund, 2003).

Ever since Ramon y Cajal's first description of local-circuit interneurons, the molecular identity and network contributions of GABAergic cells have been studied in postnatal cortical networks. It is only through recent advances of molecular genetics and cell biology that we have come to appreciate the interneurons’ different origins, migratory behaviours and cytoarchitectural features (Xu et al. 2004; Butt et al. 2005; Miyoshi et al. 2010; Tricoire et al. 2011). Although present in limited numbers, impaired development of GABAergic interneurons in the fetal cerebrum can have permanent impact on the hippocampal circuitry (Peters & Kara, 1985), precipitating neurological disorders (Di Cristo, 2007). This review discusses recent data concerned with the developmental trajectories of CCK⁺ basket cells, and emphasizes the significance of recently identified short-range intercellular guidance cues that impact the formation and function of GABAergic synapses.

The historical approach: retrospective neurochemical analysis of CCK⁺ basket cells

Do CCK⁺ basket cells exhibit unique cytoarchitectural and molecular signalling properties that distinguish them from other basket cells? This is an important question to address to gain insights regarding their circuit parameters, response patterns, as well as development and integration into hippocampal neuronal networks (Table 1). Electrophysiology studies combined with post hoc cellular neuroanatomy were the preferred approach to identify unique characteristics of basket cells. However, the advent of molecular array technologies and genetic tagging opened new horizons in the understanding of the birthplaces, migratory routes and terminal differentiation programs of various interneuron subtypes, facilitating the resolution of ambiguities regarding molecular identities.

CCK is the primary marker to classify this group of basket cells (Fig. 1). Recent findings have revealed dual pathways downstream from CCK receptor signalling in the two basket cell populations (Foldy et al. 2007). First, CCK inhibits GABA release from CCK⁺ basket cells through CCK2 receptors. This mechanism is dependent on type 1 cannabinoid receptor (CB₁R) activation (see below) but independent of GABA release from other interneurons (Lee & Soltesz, 2011). Second, CCK2 receptors on PV⁺ basket cells respond to CCK and trigger Ca²⁺ release from intracellular Ca²⁺ stores. This event leads to depolarization by the activation of a non-selective cationic conductance enhancing GABA release (Lee et al. 2011). Therefore, the net effect is to shift inhibition from CCK⁺ to PV⁺ basket cells, leaving CCK signalling poised to modulate complex signalling networks by modifying inhibitory and excitatory signals to fine-tune precise firing patterns.

Figure 1. Characteristic molecular markers of CCK⁺ interneurons.

A, a subpopulation of 5-HT_3A-EGFP⁺ cells in the hippocampus is immunoreactive for VGLUT3 and CB₁Rs. A₁, arrowheads point to VGLUT3/CB₁R⁺ processes. B, CB₁R⁺/GAD67⁺ boutons contain the 2-AG degrading enzyme MGL (arrowheads), C–C₃, CCK⁺ interneurons express 5-HT_3ARs and CB₁Rs. Arrowheads indicate CB₁R⁺ structures. Scale bars: 20 μm (A, A₁, C), 5 μm (B) and 1 μm (C₃).

An intracellular feature of hippocampal CCK⁺ basket cells is that a subset contains vasoactive intestinal peptide (VIP) (Freund & Buzsaki, 1996). CCK⁺–VIP⁺ basket cells can co-express corticotropin-releasing factor (CRF) (Kubota et al. 2011). Once released, CRF and CCK depolarize pyramidal cells through CRF-1 and CCK2 receptors, respectively (Gallopin et al. 2006). Conspicuously, CCK⁺ basket cells express vesicular glutamate transporter 3 (VGLUT3; Fig. 1) (Harkany et al. 2004; Somogyi et al. 2004). Although the precise role of VGLUT3 in basket cells remains unknown, experiments using genetic deletion of VGLUT3 indicate that glutamate co-transmission at GABA/glycinergic synapses is necessary for the developing inhibitory circuit in the auditory system (Noh et al. 2010).

CCK⁺ interneurons also exhibit a unique assembly of cell-surface receptors. These include postsynaptic α4/β2 nicotinic acetylcholine (nAChR) (Porter et al. 1999; Ferezou et al. 2002; Bell et al. 2011), ionotropic 5-HT_3A serotonin (Morales & Bloom, 1997; Vucurovic et al. 2010) and oestrogen α receptors (Hart et al. 2007), the latter being associated with clusters of synaptic vesicles in perisomatic boutons. However, the specific role of oestrogen receptors on vesicles remains unclear. Serotonin and cholinergic signalling via 5-HT_3A and α4-nAChR, respectively, elicit fast synaptic excitation of CCK⁺–VIP⁺ basket cells (Ferezou et al. 2002; Varga et al. 2009). In contrast, oestrogen α receptor signalling limits presynaptic neurotransmitter release by reducing the likelihood of GABA-laden synaptic vesicle docking (Hart et al. 2007).

CCK⁺ interneurons express high levels of the CB₁R (Katona et al. 1999; Marsicano & Lutz, 1999; Tsou et al. 1999). CB₁Rs, a primarily G_i/o protein-coupled GPCR receptor subclass, are named after their ability to bind to and transduce the psychotropic effect of Δ⁹-tetrahydrocannabinol (THC), the major phytocannabinoid from cannabis (Devane et al. 1988; Matsuda et al. 1990). The physiological impact of agonist activation of CB₁Rs is profound since these receptors are targeted to the presynaptic terminals of many neurons (including CCK⁺ basket cells; Fig. 1) where they generally limit neurotransmitter release (Wilson & Nicoll, 2001; Kano et al. 2009; Regehr et al. 2009). CB₁R activation under physiological conditions is achieved through the ‘on-demand’ liberation of endogenous cannabinoids (‘endocannabinoids’), such as 2-arachidonoylglycerol (2-AG) (Mechoulam et al. 1995) and anandamide (Devane et al. 1992), from postsynaptic neurons. Endocannabinoid signalling belongs to the emerging family of signalling systems that mediate feedback control of neurotransmitter release through retrograde action: for 2-AG, ligand synthesis at the postsynapse occurs through diacylgycerol lipases α/β (DAGLα/β) in pyramidal cells (Bisogno et al. 2003). However, CCK⁺ interneurons lack DAGLα/βin vivo during development and adulthood identifying endocannabinoids as target-derived cues (Yoshida et al. 2011). Having traversed the synaptic cleft, 2-AG engages presynaptic CB₁Rs. Endocannabinoid inactivation operates both at postsynaptic and presynaptic loci through the segregated action of the serine hydrolase ABHD6 (Marrs et al. 2010) and monoacylglycerol lipase (MGL) (Dinh et al. 2002), respectively. CCK⁺ basket cells express and co-target MGL with CB₁Rs to their presynapses (Gulyas et al. 2004; Yoshida et al. 2011). Furthermore, cyclooxygenase-2 (COX-2) has been demonstrated to degrade 2-AG (Kozak et al. 2000) and its inhibition prolongs depolarization-induced suppression of inhibition (DSI) in a CB₁R-dependent manner affirming that COX-2 is involved in CB₁R-mediated retrograde signalling (Kim & Alger, 2004; Straiker & Mackie, 2009) (Fig. 3).

Figure 3. Enzyme-dependent divergent time courses for DSI- and 2-AG-mediated inhibition in DSI_fast- and DSI_slow-expressing cultured interneurons.

A, duration of DSI (3 s depolarization) in DSI_fast (red) and DSI_slow (blue) neurons, as a result of MGL acting alone (DSI_slow) or in combination with COX-2 (DSI_fast). Aa, bar graph shows t_1/2 values with 99% confidence intervals. B, duration of 2-AG-mediated inhibition in DSI_fastvs. DSI_slow neurons. Ba, bar graph shows t_1/2 values with 99% confidence intervals. *99% confidence intervals do not overlap. Reprinted from Neuroscience163, Straiker & Mackie (2009), Cannabinoid signaling in inhibitory autaptic hippocampal neurons, pp. 190–201, Copyright 2009, with permission from Elsevier.

CCK⁺ basket cells also express presynaptic GABA_B receptors (Sloviter et al. 1999). Although the role of GABA_B receptors remains largely unclear, a coupling to N-type Ca²⁺ channels (Wilson et al. 2001) in a Ca²⁺-dependent manner similar to CB₁Rs (Neu et al. 2007) has been reported.

Should I stay or should I go: the developmental program of CCK interneurons

Understanding the mechanisms driving the development of any neuron and its network requires detailed knowledge about the genetic and environmental history of the cell. This is particularly challenging for telencephalic interneurons since it is impossible to deduce the precise birth location of a neuron based on where it is found in the adult structure. Despite these challenges, the origins and genetic programs involved in the developmental control of neocortical interneuron differentiation are beginning to be understood (Batista-Brito & Fishell, 2009). In contrast to pyramidal cells, interneurons migrate long distances tangentially to reach their final destinations by and after birth (Fig. 2). The medial ganglionic eminence (MGE) was long thought to give rise to around 85–90% of all interneurons (Nery et al. 2002). However, recent data suggest that as many as 30–40% of cortical interneurons are instead derived from the caudal ganglionic eminence (CGE, Fig. 2) (Lee et al. 2010a; Miyoshi et al. 2010). The phenotypes of CGE-derived interneurons are non-overlapping with their MGE-derived counterparts, and these neurons are generally born later during development (Butt et al. 2005; Miyoshi et al. 2010; Tricoire et al. 2011).

Figure 2. Molecular cues orchestrating CCK⁺ basket cell development.

Overview of the molecular cues involved in the migration and differentiation of basket cells towards the neocortex and the hippocampus. A and B, basket cells born in the CGE migrate through the marginal zone (MZ) or the intermediate zone (IZ) of the cerebral cortex (Manent et al. 2005; Morozov et al. 2009) to find their final positions either in the neocortex (A) or the hippocampus (B). Green trajectories indicate the migratory routes of prospective GABA interneurons. Molecular cues for each developmental state are noted. Question marks query the specificity of cues during CCK⁺ interneuron development.

CCK⁺ basket cells are CGE derived

All cortical CGE-derived interneurons express 5-HT_3A receptors, as well as respond to nicotinergic stimulation (Lee et al. 2010a). Therefore, the use of a 5-HT_3ABAC-EGFP reporter mouse line allowed detailed analyses showing that all VIP⁺ cells are born in the CGE (Lee et al. 2010a; Vucurovic et al. 2010). Some of these deep layer VIP cells were shown to co-express CCK. A recent genetic fate mapping study of the hippocampus using GAD65-EGFP and Mash1CreER mice substantiated these findings by showing that CGE-derived cells included most CCK⁺ interneurons (Tricoire et al. 2011). However, a definitive conclusion about the origin of the entire cortical/hippocampal CCK population remains elusive for a number of reasons: (1) the GAD65-EGFP and Mash1CreER transgenic models suffer from an incomplete labelling of the entire CGE-derived neuron population (Miyoshi et al. 2010; Tricoire et al. 2011). (2) The 5-HT_3ABAC-EGFP mouse has not yet been shown to be as specific for CGE-derived cells in the hippocampus as it is in the cortex. (3) In contrast to available immunohistochemical data, a stringent genetic design found a higher number of GFP-tagged cells in superficial cortical laminae, when crossing CCK-IRES-Cre (Taniguchi et al. 2011) with a Dlx5/6-Flpe and a RCE-dual reporter (Miyoshi et al. 2010), which expresses EGFP only upon coincident Flpe- and Cre-driven recombination. This suggests that either VIP⁻ CCK cells escaped detection in earlier studies or there is a population of interneurons transiently expressing CCK during development.

Genetic regulation of interneuron specification

A number of transcription factors are crucial for general cortical interneuron development. These include the Dlx gene family and the proneural gene Mash1 (Long et al. 2009). Mice lacking both Dlx1 and Dlx2 have a severe loss of tangentially migrating interneurons (Cobos et al. 2007). Similarly, Mash1 null mice have a marked loss of GABAergic cells in the neocortex and hippocampus (Anderson et al. 1997; Casarosa et al. 1999). The specific genetic program that controls the specification and generation of interneurons derived from the MGE, and involving the sequential activation of Nkx2-1 (initial specification) followed by Lhx6 and Sox6 (migration and differentiation) is beginning to be elucidated (Liodis et al. 2007; Du et al. 2008; Azim et al. 2009; Batista-Brito et al. 2009).

The search is still on for a CGE-specific factor equivalent to Nkx2-1. Such a transcription factor is probably repressed by Nkx2-1 since removal of Nkx2-1 after the final cell division of interneuron progenitors dictates the acquisition of a full CGE phenotype in many MGE-derived cells (Butt et al. 2008). One gene family that is preferentially expressed in the CGE during development includes the Coup-TF1/2 genes (Kanatani et al. 2008). Accordingly, Coup-TF1 removal specifically in interneurons reduces the amount of CGE-derived VIP⁺ and CR⁺ interneurons (Lodato et al. 2011). Coup-TF2 has also been implicated in regulating tangential migration towards the cerebral cortex, and is expressed in a subset of CB₁R⁺, as well as CB₁R⁻, hippocampal cells (Kanatani et al. 2008; Fuentealba et al. 2010; Antypa et al. 2011). Although the hunt for a CGE-specific ‘master regulator’ might be rewarding, the hypothesis that CGE fate may be a ‘default state’ of interneurons derived from the ganglionic eminences also seems plausible.

Developmental cues for CCK⁺ basket cells

Although cell-autonomous mechanisms can drive the initial engagement of CGE-derived basket cells in migratory behaviours and cytoarchitectural differentiation, short-range paracrine signals will provide critical cues to instruct the directionality of cell movement, synapse formation and functional integration into neuronal networks (Fig. 2).

Classical neurotransmitters impact interneuron development. In particular, serotonin has been identified as a cue instructing interneuron migration, differentiation and synaptogenesis (Lauder, 1990). Inhibition of serotonin synthesis in pregnant rats leads to a decrease in the migration and terminal differentiation of CCK⁺ interneurons in affected offspring (Vitalis et al. 2007). A gain-of-function analysis using SLC6A4 knockouts in which serotonin reuptake is blocked (‘hyperserotonergic mouse’) demonstrated an increase of CGE-derived GAD65⁺ interneurons in the cerebral cortex (Riccio et al. 2009). Data from organotypic slice systems suggest that serotonin's concentration is critical to define the directionality of its action: excessive bath-applied serotonin decreased interneuron migration. This effect was mediated by 5-HT₆ but not 5-HT_3A receptors (Riccio et al. 2009). Although its direct impact on CCK⁺ interneurons is less well understood, dopamine D1 receptor activation promotes interneuron migration from both the MGE and CGE. In contrast, dopamine D2 receptors (D2Rs) limit the tangential migration of interneurons (Crandall et al. 2007). Similarly, disrupted dopamine signalling by either inhibition of tyrosine hydroxylase or stimulation of D2Rs limits the size of the Dlx5a/6aIG-GFP⁺ GABAergic interneuron pool in zebrafish (Souza et al. 2011).

GABA signalling is thought to be excitatory during embryogenesis and early postnatal development (Ben-Ari, 2002). The conversion to an inhibitory mode of GABA action relies on the coincident and opposite expressional regulation of the neuron-specific K⁺–Cl⁻ co-transporter (KCC2, increase) and the Na⁺–K⁺–Cl⁻ co-transporter (NKCC1, decrease), resulting in a significant decrease of the intracellular Cl⁻ concentrations (Liu et al. 2006). Manipulations that limit the loss of NKCC1 prevent the developmental switch of the Cl⁻ gradient (Liu et al. 2006). Cholinergic neurotransmission through nAChRs is needed for the developmental switch. Accordingly, α7-nAChR^−/− mice retain high NKCC1 and low KCC2 levels, precluding inhibitory GABA neurotransmission (Liu et al. 2006) associated with morphological irregularities in hippocampal neurons (Liu et al. 2007). Since CCK⁺ basket cells express both α7-nAChRs and GABA_A receptors, it is plausible to assume that nicotine stimulation (e.g. maternal tobacco smoking) could compromise the morphological differentiation – particularly synaptogenesis – of this cell type. Short-range GABA signals, acting on GABA_A receptors, are also required to maintain tangential migration through leading process elongation in vivo (Manent et al. 2005).

Neurotrophins are indispensable for interneuron development. Brain-derived neurotrophic factor (BDNF) powerfully regulates the morphological and neurochemical differentiation of GABAergic interneurons (Marty et al. 1996; Berghuis et al. 2004). GABA signalling acts upstream to BDNF by repressing BDNF synthesis (Marty et al. 1996) to decrease neurite outgrowth. Yet other signalling cassettes can hijack (‘trans-activate’) the tropomyosin-related kinase B (TrkB) receptor on interneurons in the absence of its cognate ligand. Endocannabinoids are one such class of signalling molecules using TrkB receptors to regulate CCK⁺ interneuron migration (Berghuis et al. 2005). The idea that endocannabinoid signalling via CB₁Rs is involved in interneuron development is reinforced by several findings: firstly, by the ability of THC to induce redistribution of hippocampal CCK⁺ interneurons in utero (Berghuis et al. 2005). Secondly, acute exposure of the growth cones of CB₁R⁺/CCK⁺ interneurons to endocannabinoid microgradients evokes chemorepulsive turning or collapsing responses (Berghuis et al. 2007). Thirdly, CB₁R activation inhibits neurite outgrowth, and abolishes BDNF-induced axonal growth (Berghuis et al. 2005). Cumulatively, these data suggest an antagonistic interplay between endocannabinoid and neurotrophin signalling cassettes in the refinement of interneuron morphology and synaptic wiring. Interestingly, these effects prevail once the synapse forms with remodelled endocannabinoid and BDNF signalling networks participating in negative and positive retrograde feedback loops to control synaptic efficacy, respectively.

Endocannabinoid control of synaptic plasticity

Theta oscillations in the hippocampus occur during exploration and rapid eye movement sleep and are involved in place finding and learning and memory (O’Keefe & Nadel, 1978; Buzsaki, 2002). During these oscillations, pyramidal cells transmit information. PV⁺ basket cells synchronize the rhythm of the network, while endocannabinoid-sensitive CCK⁺ basket cells function as fine-tuning devices (Freund, 2003). In anesthetized rodents, CCK⁺ interneurons fire on the ascending phase of the theta wave, at the moment when hippocampal place cells become activated (Klausberger et al. 2005). In contrast, PV⁺ basket cells fire on the descending phase (Klausberger et al. 2003). As we discussed above, CCK⁺ basket cells play a critical role in feed-forward inhibition by releasing CCK, resulting in the enhancement of GABA release from PV⁺ interneurons (Foldy et al. 2007). Their inhibition of pyramidal cells ceases upon activation of CB₁Rs on their presynaptic terminals, forming a spatial focus of activity. Coincidentally, they maintain inhibitory control over resting place cells through entrainment of PV⁺ interneurons (Carlson et al. 2002; Chevaleyre & Castillo, 2004; Klausberger et al. 2005).

Amongst all hippocampal neurons, CCK⁺ interneurons express the highest known levels of CB₁Rs (Katona et al. 1999). This gives rise to diverse mechanisms of endocannabinoid-mediated short- and long-term synaptic plasticity at their synaptic inputs onto pyramidal neurons, including depolarization-induced suppression of inhibition (DSI) (Pitler & Alger, 1992; Ohno-Shosaku et al. 2001; Wilson & Nicoll, 2002), metabotropic suppression of inhibition (MSI) (Varma et al. 2001; Kim et al. 2002) and long-term depression (iLTD) (Chevaleyre & Castillo, 2003; Chevaleyre et al. 2007).

Of these, the inhibitory perisomatic inputs deriving from CCK⁺ basket cells are the best characterized. The CCK⁺–CA1 pyramidal synapse was one of the first sites shown to express DSI (Pitler & Alger, 1992; Ohno-Shosaku et al. 2001; Wilson & Nicoll, 2002). DSI is a form of synaptic plasticity that is induced postsynaptically, but acts presynaptically via a messenger traversing the synaptic cleft in a direction opposite to that of the major neurotransmitter (‘retrograde messenger’). Accordingly, 2-AG, the likely retrograde messenger (Gao et al. 2010; Tanimura et al. 2010), is produced in a Ca²⁺-dependent manner via postsynaptic activation of DAGLα, which cleaves the DAG precursor into 2-AG. The release of lipophilic 2-AG is facilitated, yet non-vesicular, contrasting classical vesicle-dependent neurotransmitter release (Neu et al. 2007). Once liberated, 2-AG (and other endocannabinoids) activates presynaptic CB₁Rs, which subsequently reduce neurotransmitter release by inhibiting N-type Ca²⁺ channels (Wilson et al. 2001). DSI can last for tens of seconds. However, its duration is chiefly determined by the expression of one or more pre- or post-synaptic 2-AG-degrading enzymes (Kim & Alger, 2004; Hashimotodani et al. 2007; Straiker et al. 2009), which can act in concert to terminate 2-AG signalling. An example of enzymatic cooperativity is shown in Fig. 3, where the two temporally distinct forms of DSI (referred to as DSI_fast and DSI_slow) – observed in cultured neurons – are determined by the complement of MGL and COX-2 available to break down 2-AG (Straiker & Mackie, 2009).

Endocannabinoid signalling is not restricted to CCK⁺ basket cell to CA1 pyramidal neuron synapses. For instance, CCK⁺ basket cell synapses also target other CCK⁺ or PV⁺ basket cells (Karson et al. 2009). Although studying interneuron–interneuron coupling is challenging, evidence for domain-specific plasticity of outputs has begun to appear (Lee et al. 2010b), including endocannabinoid plasticity at three inputs/outputs of CCK⁺ Schaffer collateral-associated interneurons (Ali & Todorova, 2010).

Whilst dendritic spines of pyramidal cells are the consensus sites for DAGL's subcellular accumulation in the neocortex and hippocampus (Yoshida et al. 2006), synaptically connected CCK⁺ basket cells can express endocannabinoid-mediated DSI in the apparent absence of DAGLα (Daw et al. 2009). This phenomenon could be explained by a complex metabolic configuration of synaptic plasticity through the release of other endocannabinoids, such as anandamide (Ali, 2007; Puente et al. 2011), or by other forms of endocannabinoid actions like the direct potentiation of postsynaptic GABA_A receptors by 2-AG (Sigel et al. 2011). Given that at least ∼40% of cultured hippocampal interneurons express the machinery to sustain endocannabinoid signalling in response to depolarization (Straiker & Mackie, 2009), this suggests that many CA1 interneurons may be more than just targets of endocannabinoid signalling.

Perisomatic endocannabinoid-mediated plasticity by CCK⁺ basket cells is well positioned to regulate the output of CA1 pyramidal neurons. Their role may be that of an integrator of assorted weaker inputs to support feedback inhibition at key sites of pyramidal neuron output regulation (Glickfeld & Scanziani, 2006). However, endocannabinoid-mediated synaptic plasticity is itself under modulation: endocannabinoid inhibition can be overridden by a high (>20Hz) firing rate in CCK⁺ basket cells (Foldy et al. 2006). In addition, indirect evidence suggests that postsynaptic CCK receptors can enhance endocannabinoid release (Foldy et al. 2007).

Conclusions: disease implications

This review provides a concise summary of the molecular and network features that make CCK⁺ basket cells unique, and indispensable for the proper function of hippocampal neuronal networks. If a specific class of interneurons is required to maintain a form of control over the hippocampal circuitry then its loss should manifest in altered behaviours or neuropsychiatric illness.

Endocannabinoids have also been suggested to play a role in the prevention of epileptic activity, particularly in the neonatal period when they can act as a substitute for inhibitory GABA (note that GABA is excitatory during this period; Ben-Ari, 2002). In the immature hippocampus, CB₁R antagonism leads to epileptic discharges, while receptor stimulation reduces network activity (Bernard et al. 2005). However, and in accord with the role of endocannabinoids during axon guidance (Berghuis et al. 2007; Mulder et al. 2008; Keimpema et al. 2010), prenatal (‘ectopic’) stimulation of CB₁Rs by full agonists results in permanent modifications of the hippocampal circuitry such that decreased glutamatergic excitation persists in the offspring (Mereu et al. 2003). Therefore, CB₁R signalling appears unexpectedly fundamental for the proper development and function of neuronal networks.

Another compelling example of this lies with the molecular pathogenesis of schizophrenia. BDNF and TrkB expression are decreased in schizophrenic patients (Hashimoto et al. 2005). In addition, CCK levels are also reduced in schizophrenics (Bachus et al. 1997; Fung et al. 2010). Since BDNF influences the differentiation of CCK⁺ interneurons (Marty et al. 1996; Berghuis et al. 2005), the decrease in BDNF, together with a decrease in CCK content, argues for the loss of CCK⁺ interneurons. This hypothesis is supported by the coincident decrease of CB₁R and CCK mRNAs in schizophrenia (Eggan et al. 2008). It is interesting to note that these correlative changes are not a consequence of treating the psychosis but inherent to the disease itself (Bachus et al. 1997; Eggan et al. 2008). In contrast, higher CCK (Bachus et al. 1997) and CB₁R levels (Hungund et al. 2004) have been reported in suicide victims, suggesting an increased potential for modulation of GABAergic neurotransmission. These findings suggest that CCK⁺ basket cell functions underlie fundamental behavioural and cognitive traits, and that their dysfunction can precipitate devastating neuropsychiatric symptoms.

An emerging consensus is that most, if not all, neuropsychiatric disorders have developmental origins. A developmental component is particularly well characterized in schizophrenia, and centres around the misrouting of interneurons and inappropriate endocannabinoid signalling (Di Cristo, 2007). Further studies focusing on the birth, migration, differentiation and functions of defined subsets of interneurons integrating into the hippocampal circuitry will therefore be essential to drive the development of successful therapeutic interventions.

Glossary

Abbreviations

2-AG

2-arachidonoylglycerol

ABHD6

α-β hydrolase domain 6

BDNF

brain-derived neurotrophic factor

CB₁R

type 1 cannabinoid receptor

CCK

cholecystokinin

CGE

caudal ganglionic eminence

COX-2

cyclooxygenase-2

CRF

corticotropin releasing factor

ctx

cortex

D2R

dopamine receptor D2

DAGLα

diacylglycerol lipase α

DSI

depolarization-induced suppression of inhibition

GABA

γ-aminobutyric acid

hip

hippocampus

iLTD

long-term depression of inhibition

IZ

intermediate zone

LGE

lateral ganglionic eminence

MGE

medial ganglionic eminence

MGL

monoacylglycerol lipase

MSI

metabotropic suppression of inhibition

MZ

marginal zone

nAChR

nicotinic acetylcholine receptor

NPY

neuropeptide Y

PV

parvalbumin

SOM

somatostatin

TrkB

tropomyosin-related receptor kinase B

THC

tetrahydrocannabinol

VGLUT3

vesicular glutamate transporter 3

VIP

vasoactive intestinal peptide

VZ

ventricular zone